Exploring the Chemical Composition of Female Zucchini Flowers for Their Possible Use as Nutraceutical Ingredient.

The zucchini (Cucurbita pepo L.) plant is well known for its fruits; however, its edible flowers appear to contain several active molecules, including polyphenols, which display poor bioaccessibility after gastrointestinal digestion (GiD). This study explores the bioaccessibility of polyphenols and antioxidant capacity within zucchini flower extracts during simulated GiD. Two nutraceutical formulations, non-acid-resistant (NAcR) and acid-resistant (AcR) capsules containing an aqueous extract of zucchini flowers, were employed in this investigation. Additionally, high-resolution mass spectrometry (Q-Orbitrap HRMS) was utilized for a comprehensive analysis of their polyphenolic constituents. Predominantly, rutin and isorhamnetin-3-rutinoside were the most prevalent compounds detected in the samples (514.62 and 318.59 mg/kg, respectively). Following in vitro GiD, the extract encapsulated in AcR capsules exhibited enhanced bioaccessibility during both the duodenal (189.2 and 162.5 mg GAE/100 g, respectively) and colonic stages (477.4 and 344.7 mg GAE/100 g, respectively) when compared with the extract encapsulated in NAcR capsules. This suggests that gastric acidity adversely impacted the release of polyphenols from NAcR capsules. In conclusion, the aqueous zucchini flower extract emerges as a promising and readily accessible source of dietary polyphenols. Moreover, the utilization of AcR capsules presents a potential nutraceutical formulation strategy to improve polyphenol bioaccessibility, enhancing its applicability in promoting health and well-being.

Multi-Mycotoxin Method Development Using Ultra-High Liquid Chromatography with Orbitrap High-Resolution Mass Spectrometry Detection in Breakfast Cereals from the Campania Region, Italy.

Breakfast cereals have been reported as one of the most susceptible cereal-based products to mycotoxin contamination. These products pose an even more concerning risk to human health since they are marketed as a ready-to-eat product and one of its main population targets is children. Therefore, the main goal of the present study was to conduct a monitoring study of multiple mycotoxins contained in breakfast cereals samples marketed in Italy through ultra-high performance liquid chromatography coupled to high-resolution Q-Orbitrap tandem mass spectrometry. An acetonitrile-based methodology was validated for quantifying 24 mycotoxins in breakfast cereals. The results showed that 93% of the samples contained at least one mycotoxin. Beauvericin was the most prevalent toxin (86% of samples; mean concentration: 30.66 µg/kg), although the main enniatins, zearalenone-derived forms and fumonisins B1 and B2 were also detected. Co-occurrence was observed in 73% of the positive samples with up to five mycotoxins simultaneously occurring, mainly due to the combination of beauvericin and enniatins. These results provided more evidence about the high impact of non-regulated mycotoxins, such as the emerging Fusarium toxins, in breakfast cereals, and encourages the development of analytical methodologies including these and zearalenone-derived forms that could be going unnoticed with current methodologies.

Multi-Mycotoxin Analysis in Italian Grains Using Ultra-High-Performance Chromatography Coupled to Quadrupole Orbitrap Mass Spectrometry.

Mycotoxins are a major source of contamination in cereals, posing risks to human health and causing significant economic losses to the industry. A comprehensive strategy for the analysis of 21 mycotoxins in Italian cereal grain samples (n = 200) was developed using a simple and quick sample preparation method combined with ultra-high-performance liquid chromatography coupled with quadrupole Orbitrap high-resolution mass spectrometry (UHPLC Q-Orbitrap HRMS). The proposed method showed some advantages, such as multi-mycotoxin analyses with simple sample preparation, fast determination, and high sensitivity. The analysis of the sample revealed the presence of 11 mycotoxins, with α-zearalenol being the most frequently detected, while deoxynivalenol exhibited the highest contamination level. Furthermore, co-occurrence was identified in 15.5% of the samples under analysis. Among these, 13% of the samples reported the simultaneous presence of two mycotoxins, while 2.5% showed the co-occurrence of three mycotoxins. Currently, there has been a renewed interest in guaranteeing the quality and safety of products intended for human consumption. This study holds significant value due to its ability to simultaneously detect multiple mycotoxins within a complex matrix. Furthermore, it provides findings regarding the occurrence and co-occurrence of emerging mycotoxins that currently lack regulation under the existing European Commission Regulation.

Analysis of Mycotoxin and Secondary Metabolites in Commercial and Traditional Slovak Cheese Samples.

Cheese represents a dairy product extremely inclined to fungal growth and mycotoxin production. The growth of fungi belonging to Aspergillus, Penicillium, Fusarium, Claviceps, Alternaria, and Trichoderma genera in or on cheese leads to undesirable changes able to affect the quality of the final products. In the present investigation, a total of 68 types of commercial and traditional Slovak cheeses were analyzed to investigate the occurrence of fungal metabolites. Altogether, 13 fungal metabolites were identified and quantified. Aflatoxin M1, the only mycotoxin regulated in milk and dairy products, was not detected in any case. However, the presence of metabolites that have never been reported in cheeses, such as tryptophol at a maximum concentration level from 13.4 to 7930 µg/kg (average: 490 µg/kg), was recorded. Out of all detected metabolites, enniatin B represents the most frequently detected mycotoxin (0.06-0.71 µg/kg) in the analyzed samples. Attention is drawn to the lack of data on mycotoxins' origin from Slovak cheeses; in fact, this is the first reported investigation. Our results indicate the presence of fungal mycotoxin contamination for which maximum permissible levels are not established, highlighting the importance of monitoring the source and producers of contamination in order to protect consumers' health.

Analysis of Polyphenolic Compounds in Water-Based Extracts of Vicia faba L.: A Potential Innovative Source of Nutraceutical Ingredients.

The water-based extract of broad bean hulls contains several bioactive molecules, including polyphenols well-known to exert antioxidant activity, which could justify its use in nutraceutical formulations. Hence, the current investigation aimed to establish the polyphenolic profile of water-based extracts from broad bean hulls through UHPLC-Q-Orbitrap HRMS analysis. The findings highlighted that p-coumaric acid, chlorogenic acid, and epicatechin were the most common compounds found in the tested extracts, being quantified at a mean concentration of 42.1, 32.6, and 31.2 mg/100 g, respectively. Moreover, broad bean hull extracts were encapsulated into a nutraceutical formulation, after which the antioxidant properties and the bioaccessibility of phenolic compounds during the simulated gastrointestinal (GI) process were investigated and compared with the digested non-encapsulated extract. The data highlighted that following the GI process, the capsules were able to preserve active compounds from the adverse effects of digestion, resulting in a greater antioxidant capacity and polyphenol bioaccessibility in the duodenal and colonic phases, compared with the non-encapsulated extract. Our results showed that the water extract from broad bean hulls may be considered a valuable source of natural polyphenolic compounds; in addition, the use of a gastric-resistant capsule could be a suitable alternative to transport these bioactive compounds to the target tissues.

Bioaccessibility and Antioxidant Capacity of Bioactive Compounds From Various Typologies of Canned Tomatoes.

Tomato (Solanum lycopersicum L.) is one of the most consumed vegetables in the world; it contains high amounts of antioxidant phytochemicals and essential nutrients. Although it is commonly consumed fresh, more than 80% of its consumption derives from processed products. Since limited information on changes in the bioaccessibility of bioactive compounds during gastrointestinal digestion was reported, this current study aimed to monitor the antioxidant activity, total polyphenolic and carotenoid content, and bioaccessibility during in vitro gastrointestinal digestion of different typologies (n = 7) of canned tomatoes. A comprehensive evaluation of the polyphenolic profile of digested and not digested samples was ascertained by ultra-high-performance liquid chromatography combined with high-resolution Orbitrap mass spectrometry. The results highlighted a considerable content of rutin (1.191-9.516 mg/100 g), naringenin (0.359-1.452 mg/100 g), chlorogenic acid (1.857-11.236 mg/100 g), and lycopene (50.894-222.061 mg/kg) in the analyzed matrices. After in vitro gastrointestinal digestion, large variability, losses and low recovery were recorded. An appreciable percentage of rutin (30.7%), naringenin (29.6%), chlorogenic acid (25.8%), and lycopene (varied between 9.3 and 20%) remained bioaccessible after the in vitro gastrointestinal digestion. Our study could be a valid support to evaluate which content of bioactive compounds could be really bioaccessible to exercise beneficial effects on human health.

Effect of Different Coffee Brews on Tryptophan Metabolite-Induced Cytotoxicity in HT-29 Human Colon Cancer Cells.

Coffee consumption positively influences colon health. Conversely, high levels of tryptophan metabolites such as skatole released from intestinal putrefactive fermentation in the presence of excessive dietary animal protein intake, and gut microbiota alterations, may have several adverse effects, including the development of colorectal cancer. Therefore, this study aimed to elucidate the potential protective effects of coffee in the presence of different skatole levels. The results showed that skatole exposure induced reduced cell viability and oxidative stress in the HT-29 human colon cancer cell line. However, co-treatment of cells with skatole and coffee samples was able to reduce ROS production (up to 45% for espresso) compared to cells not treated with coffee. Real-time PCR analysis highlighted that treating HT-29 cells with skatole increased the levels of inflammatory cytokines and chemokines TNF-α, IL-1ß, IL-8, and IL12, whereas exposure to coffee extracts in cells that were pretreated with skatole showed anti-inflammatory effects with decreased levels of these cytokines. These findings demonstrate that coffee may counteract the adverse effects of putrefactive compounds by modulating oxidative stress and exerting anti-inflammatory activity in colonocytes, thus suggesting that coffee intake could improve health conditions in the presence of altered intestinal microbiota metabolism.

Effect of Dewaxed Coffee on Gastroesophageal Symptoms in Patients with GERD: A Randomized Pilot Study.

Gastroesophageal Reflux Disease (GERD) is multifactorial pathogenesis characterized by the abnormal reflux of stomach contents into the esophagus. Symptoms are worse after the ingestion of certain foods, such as coffee. Hence, a randomized pilot study conducted on 40 Italian subjects was assessed to verify the effect of standard (SC) and dewaxed coffee (DC) consumption on gastroesophageal reflux symptoms and quality of life in patients with gastrointestinal diseases. The assessment of patient diaries highlighted a significant percentage reduction of symptoms frequency when consuming DC and a significant increase in both heartburn-free and regurgitation-free days. Consequentially, patients had a significant increase of antacid-free days during the DC assumption. Moreover, the polyphenolic profile of coffee pods was ascertained through UHPLC-Q-Orbitrap HRMS analysis. Chlorogenic acids (CGAs) were the most abundant investigated compounds with a concentration level ranging between 7.316 (DC) and 6.721 mg/g (SC). Apart from CGAs, caffeine was quantified at a concentration level of 5.691 mg/g and 11.091 for DC and SC, respectively. While still preliminary, data obtained from the present pilot study provide promising evidence for the efficacy of DC consumption in patients with GERD. Therefore, this treatment might represent a feasible way to make coffee more digestible and better tolerated.

In Vitro Bioaccessibility and Antioxidant Activity of Polyphenolic Compounds from Spent Coffee Grounds-Enriched Cookies.

Spent coffee ground (SCG) is a significant by-product generated by the coffee industry. It is considered a great source of bioactive molecules well-recognized for exerting biological properties. This study aimed to implement SCG in a baked foods, such as cookies (SCGc), to increase their bioactive potential. A comprehensive study of the polyphenolic fraction of the SCG and SCGc using a high-resolution mass spectrometry analysis was performed. Moreover, the polyphenol bioaccessibility and change in antioxidant activity during simulated gastrointestinal digestion (GiD) were assessed. Data showed that SCGc provided 780 mg of melanoidins, 16.2 mg of chlorogenic acid (CGA), 6.5 mg of caffeine, and 0.08 mg of phenolic acids per 100 g of sample. Moreover, the 5-caffeoylquinic acid was the most relevant CGA found in SCG (116.4 mg/100 g) and SCGc (8.2 mg/100 g) samples. The antioxidant activity evaluated through three spectrophotometric tests, and the total phenolic compounds of SCGc samples exhibited significantly higher values than the control samples. Furthermore, during simulated GiD, the highest bioaccessibility of SCGc polyphenols was observed after the colonic stage, suggesting their potential advantages for human health. Therefore, SCG with high content in bioactive molecules could represent an innovative ingredient intended to fortify baked food formulations.

Chemical Composition of Green Pea (Pisum sativum L.) Pods Extracts and Their Potential Exploitation as Ingredients in Nutraceutical Formulations.

Agro-industrial wastes contain a large number of important active compounds which can justify their use as innovative ingredients in nutraceutical products. This study aimed to provide a complete analysis of active molecules, namely (poly)phenols in pea pods water-based extracts, through a UHPLC-Q-Orbitrap HRMS methodology. Data showed that 5-caffeoylquinic acid, epicatechin, and hesperidin were the most relevant (poly)phenols found in the assayed extracts, with a mean value of 59.87, 29.46, and 19.94 mg/100 g, respectively. Furthermore, changes in antioxidant capacity and bioaccessibility of total phenolic compounds (TPC) after the simulated gastrointestinal (GI) process were performed using spectrophotometric assays (FRAP, DPPH, ABTS, and TPC by Folin-Ciocalteu). The acid-resistant capsules (ARC) and the non-acid resistant capsules (NARC) containing the pea pod extract underwent simulated GI digestion. The results suggested that the ARC formulations were able to preserve the active compounds along the simulated GI process, highlighting a higher TPC value and antioxidant capacity than the NARC formulations and the not-encapsulated extracts. Hence, the pea pods water-based extracts could be utilized as a potential alternative source of active compounds, and the use of ARC could represent a suitable nutraceutical formulation to vehiculate the active compounds, protecting the chemical and bioactive properties of (poly)phenols.

Antioxidant and Anti-Inflammatory Activity of Coffee Brew Evaluated after Simulated Gastrointestinal Digestion.

Coffee contains human health-related molecules, namely polyphenols that possess a wide range of pharmacological functions, and their intake is associated with reduced colon cancer risk. This study aimed to assess the changes in the anti-inflammatory and antioxidant activity of coffee after simulated gastrointestinal digestion. The evaluation of intracellular reactive oxygen species (ROS) levels in the HT-29 human colon cancer cell line and three in vitro spectrophotometric assays were performed to determine the antioxidant activity of the samples. Characterization of coffee composition was also assessed through a Q-Orbitrap high-resolution mass spectrometry analysis. The results highlighted that the levels of polyphenols in the digested coffee brews were higher than those of the non-digested ones. All assayed samples decreased the levels of intracellular ROS when compared to untreated cells, while digested coffee samples exhibited higher antioxidant capacity and total phenolic content than not-digested coffee samples. Digested coffee samples showed a higher reduction in interleukin-6 levels than the not-digested samples in lipopolysaccharide-stimulated HT-29 cells treated for 48 h and fewer cytotoxic effects in the MTT assay. Overall, our findings suggest that coffee may exert antioxidant and anti-inflammatory properties, and the digestion process may be able to release compounds with higher bioactivity.

Female mating tactics in lekking fallow deer (Dama dama): experience explains inter-individual variability more than costs.

Most studies on ungulate reproduction have focused on the covariates of male reproductive success, while there is much less information on female tactics of mate choice. The aim of this work is to fill this gap and to assess condition-dependent variations in female tactics in a lekking fallow deer (Dama dama) population. In particular, we investigated three indirect selection mechanisms: i) aggregation: when females join an already formed female group; ii) copying: when females copy the mate choice of other females and iii) territory choice: when females select a territory where many copulations had previously occurred. Our results show that female fallow deer, which are less experienced (younger) and/or incur higher travel costs (home range far from the lek), adopt indirect forms of mate selection more often than older females or females residing near the lek, respectively. Compared to adults, younger females remained longer in the lek (almost three times) and in male territories, returning to the lek after copulation. However, despite the time spent at the lek, younger females were not able to select the highest-rank males, and relied on territory choice more often than older females. Farther does visited the lek less frequently (farthest females only once) and arrived on average 5 days later than closer females (which performed up to 7 visits), but they were seen more often within female groups (aggregation). We did not find a different amount of copying in younger or in farther females. Our results contribute to advance our understanding of female behaviours in ungulate leks.

Generalized structural equations improve sexual-selection analyses.

Sexual selection is an intense evolutionary force, which operates through competition for the access to breeding resources. There are many cases where male copulatory success is highly asymmetric, and few males are able to sire most females. Two main hypotheses were proposed to explain this asymmetry: "female choice" and "male dominance". The literature reports contrasting results. This variability may reflect actual differences among studied populations, but it may also be generated by methodological differences and statistical shortcomings in data analysis. A review of the statistical methods used so far in lek studies, shows a prevalence of Linear Models (LM) and Generalized Linear Models (GLM) which may be affected by problems in inferring cause-effect relationships; multi-collinearity among explanatory variables and erroneous handling of non-normal and non-continuous distributions of the response variable. In lek breeding, selective pressure is maximal, because large numbers of males and females congregate in small arenas. We used a dataset on lekking fallow deer (Dama dama), to contrast the methods and procedures employed so far, and we propose a novel approach based on Generalized Structural Equations Models (GSEMs). GSEMs combine the power and flexibility of both SEM and GLM in a unified modeling framework. We showed that LMs fail to identify several important predictors of male copulatory success and yields very imprecise parameter estimates. Minor variations in data transformation yield wide changes in results and the method appears unreliable. GLMs improved the analysis, but GSEMs provided better results, because the use of latent variables decreases the impact of measurement errors. Using GSEMs, we were able to test contrasting hypotheses and calculate both direct and indirect effects, and we reached a high precision of the estimates, which implies a high predictive ability. In synthesis, we recommend the use of GSEMs in studies on lekking behaviour, and we provide guidelines to implement these models.