In this work, the phytochemical profile and the biological properties of Colchicum triphyllum (an unexplored Turkish cultivar belonging to Colchicaceae) have been comprehensively investigated for the first time. Herein, we focused on the evaluation of the in vitro antioxidant and enzyme inhibitory effects of flower, tuber, and leaf extracts, obtained using different extraction methods, namely maceration (both aqueous and methanolic), infusion, and Soxhlet. Besides, the complete phenolic and alkaloid untargeted metabolomic profiling of the different extracts was investigated. In this regard, ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) allowed us to putatively annotate 285 compounds when considering the different matrix extracts, including mainly alkaloids, flavonoids, lignans, phenolic acids, and tyrosol equivalents. The most abundant polyphenols were flavonoids (119 compounds), while colchicine, demecolcine, and lumicolchicine isomers were some of the most widespread alkaloids in each extract analyzed. In addition, our findings showed that C. triphyllum tuber extracts were a superior source of both total alkaloids and total polyphenols, being on average 2.89 and 10.41 mg/g, respectively. Multivariate statistics following metabolomics allowed for the detection of those compounds most affected by the different extraction methods. Overall, C. triphyllum leaf extracts showed a strong in vitro antioxidant capacity, in terms of cupric reducing antioxidant power (CUPRAC; on average 96.45 mg Trolox Equivalents (TE)/g) and ferric reducing antioxidant power (FRAP) reducing power (on average 66.86 mg TE/g). Interestingly, each C. triphyllum methanolic extract analyzed (i.e., from tuber, leaf, and flower) was active against the tyrosinase in terms of inhibition, recording the higher values for methanolic macerated leaves (i.e., 125.78 mg kojic acid equivalent (KAE)/g). On the other hand, moderate inhibitory activities were observed against AChE and α-amylase. Strong correlations (p < 0.01) were also observed between the phytochemical profiles and the biological activities determined. Therefore, our findings highlighted, for the first time, the potential of C. triphhyllum extracts in food and pharmaceutical applications.

Cydonia oblonga Mill., normally known as the quince fruit, has been widely used in agro-food industries mainly to produce jams and jellies. However, other parts of the plants are still underutilized and not completely assessed for their nutraceutical profile. Therefore, in this work, the polyphenolic profile of C. oblonga was investigated using an untargeted metabolomics approach based on high-resolution mass spectrometry. Several compounds were identified in the different parts of the plants, including flavonoids (i.e., anthocyanins, flavones, flavan-3-ols, and flavonols), phenolic acids (both hydroxycinnamics and hydroxybenzoics), low-molecular-weight phenolics (tyrosol equivalents), lignans, and stilbenes. Overall, C. oblonga leaves showed the highest in vitro antioxidant potential, as revealed by 2,2-difenil-1-picrylhydrazyl (DPPH), 2,2'-Azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), ferric reducing antioxidant power (FRAP), and cupric ion reducing antioxidant capacity (CUPRAC) assays, being 189.5, 285.6, 158.9, and 348.8 mg Trolox Equivalent/g, respectively. The enzymes acetyl- and butyryl-cholinesterases were both inhibited by the different plant parts of C. oblonga, with stems showing the higher inhibitory potential. Interestingly, the fruit extracts were the only parts inhibiting the α-glucosidase, with a value of 1.36 mmol acarbose equivalents (ACAE)/g. On the other hand, strong tyrosinase inhibition was found for stems and leaves, being 72.11 and 68.32 mg Kojic acid Equivalent/g, respectively. Finally, a high number of significant (0.05 < p < 0.01) correlations were outlined between phenolics (mainly anthocyanins, flava-3-ols, and tyrosol equivalents) and the different biological assays. Taken together, our findings suggest a potential exploitation of C. oblonga leaves and stems for the food, pharmaceutical, and cosmetic industries.

Pseudomonas fluorescens is a psychrotrophic species associated with milk spoilage because of its lipolytic and proteolytic activities. Consequently, monitoring P. fluorescens or its antecedent activity in milk is critical to preventing quality defects of the product and minimizing food waste. Therefore, in this study, untargeted metabolomics and peptidomics were used to identify the changes in milk related to P. fluorescens activity by simulating the low-temperature conditions usually found in milk during the cold chain. Both unsupervised and supervised multivariate statistical approaches showed a clear effect caused by the P. fluorescens inoculation on milk samples. Our results showed that the levels of phosphatidylglycerophosphates and glycerophospholipids were directly related to the level of contamination. In addition, our metabolomic approach allowed us to detect lipid and protein degradation products that were directly correlated with the degradative metabolism of P. fluorescens. Peptidomics corroborated the proteolytic propensity of P. fluorescens-contaminated milk, but with lower sensitivity. The results obtained from this study provide insights into the alterations related to P. fluorescens 39 contamination, both pre and post heat treatment. This approach could represent a potential tool to retrospectively understand the actual quality of milk under cold chain storage conditions, either before or after heat treatments.

In this work, the comprehensive metabolomic changes in pork burgers treated with different antioxidants, namely, (a) a control without antioxidants, (b) 200 mg/kg butylated hydroxytoluene (BHT), and (c) 250 mg/kg pitanga leaf extract (PLE, from Eugenia uniflora L.), each one packaged under modified atmosphere (80% O2 and 20% CO2) for 18 days storage at 2 ± 1 °C, were deeply studied. In particular, untargeted metabolomics was used to evaluate the impact of the antioxidant extracts on meat quality. The PLE phytochemical profile revealed a wide variety of antioxidant compounds, such as polyphenols, alkaloids, and terpenoids. Multivariate statistics (both unsupervised and supervised) allowed to observe marked differences in BHT and PLE burgers metabolomic profiles during storage. Most of the differences could be attributed to hexanoylcarnitine, 4-hydroxy-2-nonenal, 6-hydroxypentadecanedioic acid, 9S,11S,15S,20-tetrahydroxy-5Z,13E-prostadienoic acid (20-hydroxy-PGF2a), sativic acid, followed by glycerophospholipids. In addition, significant correlations (p < 0.01) were observed between thiobarbituric acid reactive substances and metabolites related to lipid oxidation processes. Therefore, the approach used showed a clear modulation of lipid oxidation, likely promoted by the plant leaf extract, thus confirming the ability of PLE to delay lipid oxidative phenomena during storage.

The chemical composition of milk can be significantly affected by different factors across the dairy supply chain, including primary production practices. Among the latter, the feeding system could drive the nutritional value and technological properties of milk and dairy products. Therefore, in this work, a combined foodomics approach based on both untargeted metabolomics and metagenomics was used to shed light onto the impact of feeding systems (i.e., hay vs. a mixed ration based on hay and fresh forage) on the chemical profile of raw milk for the production of hard cheese. In particular, ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC-QTOF) was used to investigate the chemical profile of raw milk (n = 46) collected from dairy herds located in the Po River Valley (Italy) and considering different feeding systems. Overall, a total of 3320 molecular features were putatively annotated across samples, corresponding to 734 unique compound structures, with significant differences (p < 0.05) between the two feeding regimens under investigation. Additionally, supervised multivariate statistics following metabolomics-based analysis allowed us to clearly discriminate raw milk samples according to the feeding systems, also extrapolating the most discriminant metabolites. Interestingly, 10 compounds were able to strongly explain the differences as imposed by the addition of forage in the cows' diet, being mainly glycerophospholipids (i.e., lysophosphatidylethanolamines, lysophosphatidylcholines, and phosphatidylcholines), followed by 5-(3',4'-Dihydroxyphenyl)-gamma-valerolactone-4'-O-glucuronide, 5a-androstan-3a,17b-diol disulfuric acid, and N-stearoyl glycine. The markers identified included both feed-derived (such as phenolic metabolites) and animal-derived compounds (such as lipids and derivatives). Finally, although characterized by a lower prediction ability, the metagenomic profile was found to be significantly correlated to some milk metabolites, with Staphylococcaceae, Pseudomonadaceae, and Dermabacteraceae establishing a higher number of significant correlations with the discriminant metabolites. Therefore, taken together, our preliminary results provide a comprehensive foodomic picture of raw milk samples from different feeding regimens, thus supporting further ad hoc studies investigating the metabolomic and metagenomic changes of milk in all processing conditions.

Fibigia clypeata (L.) Medik. is a poorly studied plant species belonging to the Brassicaceae family, and usually used as cress in the salads. The current investigation aimed at assessing the antioxidant potential and inhibitory activity of ethyl acetate, methanol, and aqueous extracts of F. clypeata against key enzymes targeted in the management of type II diabetes (α-amylase and α-glucosidase), Alzheimer's disease (acetylcholinesterase and butyrylcholinesterase), and skin hyperpigmentation (tyrosinase). Cytotoxicity of the extracts was also determined using normal VERO and cancer FaDu and SCC-25 cell lines. Besides, LC-MS was employed to investigate the detailed phytochemical profiles of the extracts. The methanol extract showed potent enzyme inhibitory activity (4.87 mg galantamine equivalent/g, 3.52 mg galantamine equivalent/g, 126.80 mg kojic acid equivalent/g, and 24.68 mg acarbose equivalent/g, for acetylcholinesterase, butyrylcholinesterase, tyrosinase, and α-glucosidase, respectively) and antioxidant potential (96.52, 109.10, 154.02, and 104.85 mg trolox equivalent/g, for DPPH, ABTS, CUPRAC, and FRAP assays, respectively). Interestingly, caffeic acid-O-hexoside derivative, caffeyl alcohol O-glucopyranoside, and ferulic acid derivative were identified in all extracts. F. clypeata extracts showed no cytotoxicity towards VERO cell line and a weak cytotoxic potential against FaDu and SCC-25 cell lines. Interesting scientific evidence gathered from the present study support further investigation on F. clypeata in the view of designing and developing a novel therapeutic agent for the management of Alzheimer's disease, type II diabetes, skin hyperpigmentation problems, as well as cancer.

In this work, a retrospective screening based on ultra-high-performance liquid chromatography (UHPLC) coupled with high-resolution mass spectrometry (HRMS) based on Orbitrap-Q-Exactive Focus™ was used to check the occurrence of regulated and emerging mycotoxins in bulk milk samples. Milk samples were collected from dairy farms in which corn silage was the main ingredient of the feeding system. The 45 bulk milk samples were previously analyzed for a detailed untargeted metabolomic profiling and classified into five clusters according to the corn silage contamination profile, namely: (1) low levels of Aspergillus- and Penicillium-mycotoxins; (2) low levels of fumonisins and other Fusarium-mycotoxins; (3) high levels of Aspergillus-mycotoxins; (4) high levels of non-regulated Fusarium-mycotoxins; (5) high levels of fumonisins and their metabolites. Multivariate statistics based on both unsupervised and supervised analyses were used to evaluate the significant fold-change variations of the main groups of mycotoxins detected when comparing milk samples from clusters 3, 4, and 5 (high contamination levels of the corn silages) with cluster 1 and 2 (low contamination levels of the corn silages). Overall, 14 compounds showed a significant prediction ability, with antibiotic Y (VIP score = 2.579), bikaverin (VIP score = 1.975) and fumonisin B2 (VIP score = 1.846) being the best markers. The k-means clustering combined with supervised statistics showed two discriminant groups of milk samples, thus revealing a hierarchically higher impact of the whole feeding system (rather than the only corn silages) together with other factors of variability on the final mycotoxin contamination profile. Among the discriminant metabolites we found some Fusarium mycotoxins, together with the tetrapeptide tentoxin (an Alternaria toxin), the α-zearalenol (a catabolite of zearalenone), mycophenolic acid and apicidin. These preliminary findings provide new insights into the potential role of UHPLC-HRMS to evaluate the contamination profile and the safety of raw milk to produce hard cheese.

Multiple studies demonstrated biological activities of aged black garlic, including anti-inflammatory, antioxidant, and cardioprotective effects. We aimed to investigate the protective effects of an aged black garlic water extract (ABGE) alone or in association with multivitamins consisting of combined Vitamins D, C, and B12, on mouse heart specimens exposed to E. coli lipopolysaccharide (LPS). Moreover, we studied the hydrogen sulphide (H2S) releasing properties and the membrane hyperpolarization effect of the Formulation composed by ABGE and multivitamins, using Human Aortic Smooth Muscle Cells (HASMCs). ABGE, vitamins D and C, and the Formulation suppressed LPS-induced gene expression of cyclooxygenase (COX)-2, tumor necrosis factor (TNF)-α, interleukin (IL)-6, nuclear factor-kB (NF-kB), and inducible nitric oxide synthase (iNOS) on mouse heart specimens. The beneficial effects induced by the extract could be related to the pattern of polyphenolic composition, with particular regard to gallic acid and catechin. The Formulation also increased fluorescence values compared to the vehicle, and it caused a significant membrane hyperpolarization of HASMCs compared to ABGE. To conclude, our present findings showed that ABGE, alone and in association with multivitamins, exhibited protective effects on mouse heart. Moreover, the Formulation increased intracellular H2S formation, further suggesting its potential use on cardiovascular disease.

Considering that functional components of plant foods are mainly secondary-metabolism products, we investigated the shaping of health-promoting compounds in hydroponically grown butterhead lettuce (Lactuca sativa L. var. capitata) as a function of the strength of the nutrient solution utilized. To this aim, untargeted metabolomics profiling, in vitro antioxidant capacity (total phenolics, 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), cupric reducing antioxidant capacity (CUPRAC), and ferric reducing antioxidant power (FRAP) assays), and the inhibition of selected enzyme activities were investigated in two butterhead lettuce cultivars with different pigmentation, i.e., green and red Salanova. Full-strength nutrition, together with half- and quarter-strength solutions of macronutrients, was tested. Our results indicate that by reducing the nutrients strength, we could elicit a distinctive shaping of the phenolic profile of lettuce. It is noteworthy that only specific classes of phenolics (namely, lignans and phenolic acids, followed by flavones and anthocyanins) were modulated by the induction of nutritional eustress (fold-change values in the range between -5 and +11). This indicates that specific responses, rather than a generalized induction of phenolic compounds, could be observed. Nonetheless, a genotype-dependent response could be observed, with the red cultivar being much more responsive to nutritional deprivation than the green Salanova lettuce. Indeed, analysis of variance (ANOVA) confirmed a genotype x nutrition interaction in red Salanova (p < 0.001). As a consequence of the changes in phenolic composition, also the antioxidant capacity (p < 0.001) and amylase inhibition (p < 0.001) properties were affected by the growing conditions. However, the effect on cholinesterase and tyrosinase inhibition was poorly affected by the nutritional strength. Provided that yields are not compromised, the application of a controlled nutritional eustress in hydroponically cultivated lettuce may represent a valuable strategy to produce food with tailored functional features in a sustainable manner.

A carotenoid-rich extract from Lycium barbarum L. was added to extra virgin olive oil (EVOO), obtaining a carotenoid-enriched oil (EVOOCar). The oxidative stability of EVOO and EVOOCar was evaluated during long-term storage of 28 weeks at room temperature, by measuring some classical parameters (acidity and peroxide values, spectrophotometric coefficients, fatty acid composition) and the content of minor compounds (i.e., α-tocopherol and lutein). At the end of the storage, higher content (p < 0.01) of α-tocopherol in EVOOCar in respect to EVOO were observed. Zeaxanthin dipalmitate, the most abundant carotenoid compound of Goji berries, decreased slightly (p < 0.05) in EVOOCar until the end of the storage. In regard to polyphenols, an ultra-high-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (UHPLC/QTOF-MS) using untargeted metabolomics was carried out. This latter approach discriminated the two oil samples during long-term storage, allowing to identify also the phenolic classes most exposed to significant variations during storage (i.e., mainly lignans and flavones). Besides, the addition of Goji carotenoids preserved the stability of tyrosol equivalents in EVOOCar during long-term storage. These results highlighted that the enrichment of EVOO with a carotenoid-rich extract can improve the shelf-life and nutritional value of added-oil, protecting EVOO natural antioxidants during long-term storage.

This work investigated the microbiological quality and chemical profiles of two different dairy creams obtained by centrifugation vs. natural creaming separation systems. To this aim, an untargeted metabolomics approach based on UHPLC-QTOF mass spectrometry was used in combination with multivariate statistical tools to find potential marker compounds of the two different types of two dairy creams. Thereafter, we evaluated the chemical, microbiological and sensorial changes of a ricotta cheese made with a 30% milk cream (i.e., made by combining dairy creams from centrifugation and natural creaming separation) during its shelf-life period (12 days). Overall, microbiological analysis revealed no significant differences between the two types of dairy creams. On the contrary, the trend observed in the growth of degradative bacteria in ricotta during shelf-life was significant. Metabolomics revealed that triacylglycerols and phospholipids showed significant strong down-accumulation trends when comparing samples from the centrifugation and natural creaming separation methods. Additionally, 2,3-Pentanedione was among the best discriminant compounds characterising the shelf-life period of ricotta cheese (VIP score = 1.02), mainly related to sensorial descriptors, such as buttery and cheesy. Multivariate statistics showed a clear impact of the shelf-life period on the ricotta cheese, revealing 139 potential marker compounds (mainly included in amino acids and lipids). Therefore, the approach used showed the potential of a combined metabolomic, microbiological and sensory approach to discriminate ricotta cheese during the shelf-life period.

Inflammatory bowel diseases (IBDs) are chronic and multifactorial inflammatory conditions of the colonic mucosa (ulcerative colitis), characterized by increased and unbalanced immune response to external stimuli. Garlic and its bioactive constituents were reported to exert various biological effects, including anti-inflammatory, antioxidant and immunomodulatory activities. We aimed to evaluate the protective effects of a hydroalcoholic (GHE) and a water (GWE) extract from a Sicilian variety of garlic, known as Nubia red garlic, on an ex vivo experimental model of ulcerative colitis, involving isolated LPS-treated mouse colon specimens. Both extracts were able to counteract LPS-induced cyclooxygenase (COX)-2, tumor necrosis factor (TNF)-α, nuclear factor-kB (NF-kB), and interleukin (IL)-6 gene expression in mouse colon. Moreover, the same extracts inhibited prostaglandin (PG)E2, 8-iso-PGF2α, and increased the 5-hydroxyindoleacetic acid/serotonin ratio following treatment with LPS. In particular, GHE showed a better anti-inflammatory profile. The anti-inflammatory and antioxidant effects induced by both extracts could be related, at least partially, to their polyphenolic composition, with particular regards to catechin. Concluding, our results showed that GHE and GWE exhibited protective effects in colon, thus suggesting their potential use in the prevention and management of ulcerative colitis.