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A significant quantity of bone-rich poultry by-products must be disposed of by poultry processors. These products still contain a significant amount of nutritionally valuable animal proteins. In the present work, a hydrolysis protocol was optimized to recover the protein fraction of bone-rich poultry by-products while simultaneously minimizing the amount of water required for hydrolysis (thus reducing drying costs) and recycling the hydrolytic broth up to 3 times, to reduce the cost of the proteolytic enzyme. The final hydrolysis conditions involved the use of (protease from B. licheniformis, ≥2.4 U/g; 0.5 V/w of raw material) and a hydrolysis time of 2 h at 65°C. The protein hydrolysate obtained has a high protein content (79-86%), a good amino acid profile (chemical amino acid score equal to 0.7-0.8) and good gastric digestibility (about 30% of peptide bonds are already hydrolyzed before digestion). This supports its use as an ingredient in food, pet food or animal feed formulations., Competing Interests: DISCLOSURES The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

D-amino acids can affect the action of digestive enzymes, hence the protein digestion. In this work the behaviour of the main stomach and gut digestive enzymes (pepsin, trypsin, and chymotrypsin) in the presence of D-amino acids in the protein chain was monitored over time using a model peptide, Ac-LDAQSAPLRVYVE-NH 2 (belonging to β-lactoglobulin, position 48-60), where L-amino acids were systematically substituted by D-amino acids. The results showed several changes in the behaviour of digestive enzymes, not only when the D-amino acids are inserted at the specific cleavage sites (after Val-57), but in some cases also when in distant positions. The effect seemed more pronounced in the case of pepsin rather than the gut enzymes, possibly indicating a better resilience of the upper gut phase of digestion to racemization. These results demonstrated that racemization could impair nutritional value by slowing down digestibility and has different effects according to the enzyme/amino acids involved., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Nothing declared., (Copyright © 2024. Published by Elsevier Ltd.)

In this first work, commercial steak-like ( n = 3) and cured meat ( n = 3) analogues with different legume and cereal formulations were studied and compared to their animal-based ( n = 3) counterparts. Plant-based products showed lower protein content than meat controls but a good amino acidic profile even though the sum of essential amino acids of plant-cured meats does not fulfill the requirements set by the Food and Agriculture Organization for children. A comparable release of soluble proteins and peptides in the digestates after in vitro digestion was observed in meat analogues as meat products, whereas the digestibility of proteins was lower in plant-based steaks and higher in plant-based cured meats than their counterparts. The overall protein quality and digestibility of products are related to both the use of good blending of protein sources and processes applied to produce them. An adequate substitution of meat with its analogues depends mostly on the quality of raw materials used, which should be communicated to consumers.

Nowadays, consumers are increasingly inclined toward plant-based meat analogues for sake of food security, safety, and sustainability. This growing interest, not only from consumers but also from food companies, brought the offer on the market to be wide and vast. From our previous study it emerged that the market supply, especially the Italian one, is diversified both in terms of protein sources and nutrient content. Although these products are increasingly consumed, for most of the meat analogues today on the market, little is still known about their actual protein quality and digestibility. To fill this gap, in this study different commercial plant-based burgers (2 soy-based and 2 pea-based) were selected and compared to two beef burgers, as controls, in terms of protein quality and digestibility. The findings of this study demonstrated the essential amino acidic profile lacks lysine for almost all burgers (including the meat-based ones) compared to the amino acid scoring pattern set by FAO/WHO (for older children and adults), even if the sum of essential amino acids was within the range of sufficiency. All samples showed good initial protein integrity with low hydrolysis (above 6%) and percentage of D-enantiomers (above 15%). The study of the digestibility, performed by the validated INFOGEST in vitro model, showed better protein solubilisation in the case of meat burgers (63 ± 3% and 61 ± 8%), but a good digestibility also in the case of plant-based ones (from 55% to 40%). The degree of hydrolysis of the solubilised proteins was very high in all samples (from 65% to 40%) indicating a very good protein accessibility to digestive enzymes. The analysis of the peptide fraction of digestates indicated a high prevalence of collagen proteins in beef burgers and of reserve proteins in plant-based burgers. This study showed that the differences between these products are mostly dependent on the quality of the raw materials used, rather than on the vegetal or animal protein source. Therefore, to have a product with a good protein quality and digestibility, independently from the protein origin, the consumer needs to make an accurate choice, carefully reading the ingredient list., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Flavonoids are largely present in plant food such as cocoa and derived products. These compounds can interact with proteins inherently contained in the food matrix and/or the proteolytic enzymes involved in gastrointestinal digestion. The flavonoid/protein interaction might hamper protein bioaccessibility and digestibility, affecting the nutritional quality. However, information on the digestion fate of proteins in food matrices containing both proteins and flavonoids is limited. The aim of this work was to evaluate the interaction between proteins and flavonoids and verify the potential effects of this interaction on protein digestibility. Taking milk chocolate as model, first a simple whey proteins/catechins mixed system was evaluated, and then the effects on digestibility were also verified in a real sample of commercial milk chocolate. The effects of the catechins/whey proteins interaction in the model system were evaluated by optical and chiro-optical spectroscopy, outlining a slight protein structure modification upon interaction with catechins. The digestibility of the protein fraction both in the model system, with and without catechins, and also in milk chocolate, was then determined by the application of INFOGEST in vitro digestion method: the bioaccessibility was evaluated in terms of protein hydrolysis and protein solubilisation, and major peptides generated by the digestion were also determined by LC/HR-MS. Despite the slight interaction with proteins, flavonoids were found to not hinder nor modify protein solubilization, protein hydrolysis and peptide profile by digestive enzymes. Also protein digestibility in milk chocolate, evaluated by SDS-PAGE, was found to be complete. The present data clearly indicate that the interaction of the proteins with the flavonoids present in the cocoa matrix does not to affect protein bioaccessibility during digestion., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Microalgae are considered a valuable source of proteins that are used to enhance the nutritional value of foods. In this study, a standard vegetable cream recipe was reformulated through the addition of single-cell ingredients from Arthrospira platensis (spirulina), Chlorella vulgaris , Tetraselmis chui , or Nannochloropsis oceanica at two levels of addition (1.5% and 3.0%). The impact of microalgae species and an addition level on the amino acid profile and protein in vitro digestibility of the vegetable creams was investigated. The addition of microalgae to vegetable creams improved the protein content and the amino acid nutritional profile of vegetable creams, whereas no significant differences were observed in protein digestibility, regardless of the species and level of addition, indicating a similar degree of protein digestibility in microalgae species despite differences in their protein content and amino acid profile. This study indicates that the incorporation of microalgae is a feasible strategy to increase the protein content and nutritional quality of foods.

The greater awareness of consumers regarding the sustainability of food chains has shifted part of the consumption from animal protein sources to vegetable sources. Among these, of relevance both for human food use and for animal feed, is soy. However, its high protein content is unfortunately accompanied by the presence of antinutritional factors, including Kunitz's trypsin inhibitor (KTI). Now there are few analytical methods available for its direct quantification, as the inhibitory activity against trypsin is generically measured, which however can be given by many other molecules and undergo numerous interferences. Therefore, in this work, a direct label-free liquid chromatography-mass spectrometry (LC-MS) method for the identification and quantification of trypsin Kunitz inhibitor KTI3 in soybean and derivative products has been developed. The method is based on the identification and quantification of a marker peptide, specific for the protein of interest. Quantification is achieved with an external calibration curve in the matrix, and the limit of detection and the limit of quantification of the method are 0.75 and 2.51 μg/g, respectively. The results of the LC-MS method were also compared with trypsin inhibition measured spectrophotometrically, highlighting the complementarity of these two different pieces of information.

The beneficial effects of an insect-based diet on human health and, in particular, the regulatory ability of digested insects' proteins on the glycaemic response in humans are topics that need to be investigated deeper. In this work, we performed an in vitro study on the modulatory activity of gastrointestinal digested black soldier fly (BSF) prepupae on the enterohormone GLP-1 and its natural inhibitor, DPP-IV. We verified whether actions intended to valorise the starting insect biomass, i.e., insect-optimised growth substrates and prior fermentation, can positively impact human health. Our results highlight that the digested BSF proteins from all the prepupae samples had a high stimulatory and inhibitory ability on the GLP-1 secretion and the DPP-IV enzyme in the human GLUTag cell line. Gastrointestinal digestion significantly improved the DPP-IV inhibitory capacity of the whole insect protein. Moreover, it was seen that optimised diets or fermentation processes preceding the digestion, in any case, did not positively affect the efficacy of the answer. BSF was already considered one of the edible insects more suitable for human consumption for its optimal nutritional profile. The BSF bioactivity here shown, after simulated digestion, on glycaemic control systems makes this species even more promising.

Background: In Italy, there is scant evidence on the impact of Community Hospitals (CHs) on clinical outcomes., Aims: To assess the effectiveness of CHs versus long-term care hospital or inpatient rehabilitation facilities on mortality, re-admission, institutionalization, and activation of a home care programme in the Emilia-Romagna Region (ERR-Italy) after acute hospitalisation., Methods: We implemented a cohort study drawing upon the ERR Administrative Healthcare Database System and including hospital episodes of ERR residents subject ≥ 65 years, discharged from a public or private hospital with a medical diagnosis to a CH or to usual care between 2017 and 2019. To control for confounding, we applied a propensity score matching., Results: Patients transferred to CHs had a significantly lower risk of dying but an increased risk of being readmitted to community or acute hospital within 30/90 days from discharge. The hazard of institutionalisation within 30/90 days was significantly lower in the whole population of the CH exposed group but not among patients with cardiac or respiratory chronic diseases or diabetes. The activation of a home care program within 90 days was slightly higher for those who were transferred to a CH., Discussion: The findings of our study show mixed effects on outcomes of patients transferred to CHs compared to those who followed the post-acute usual care and should be taken with cautious as could be affected by the so-called 'confounding by indication'., Conclusions: The study contributes to the intermediate care available evidence from a region with a well-established care provision through CHs., (© 2022. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

The interest in agri-food residues and their valorization has grown considerably, and many of them are today considered to be valuable, under-exploited sources of different compounds and notably proteins. Despite the beneficial properties of legumes by-products, there are also some emerging risks to consider, including their potential allergenicity. In this work the immunoreactivity of chickpea, pea, and white bean by-products was assessed, and whether the production of enzymatic hydrolysates can be an effective strategy to reduce this allergenic potential. The results presented clearly indicate that the efficiency of this strategy is strongly related to the enzyme used and the food matrix. All legume by-products showed immunoreactivity towards serum of legume-allergic patients. Hydrolysates from alcalase did not show residual immunoreactivity for chickpea and green pea, whereas hydrolysates from papain still presented some immunoreactivity. However, for white beans, the presence of antinutritional factors prevented a complete hydrolysis, yielding a residual immunoreactivity even after enzymatic hydrolysis with alcalase., (© 2022. The Author(s).)

In the food and feed industry, protein extraction is commonly performed under acid or basic conditions, combined with heat, in order to increase the extraction yield. Under severe processing conditions, proteins may undergo molecular modifications. Here, the effects of heating (30, 60, 90 °C) at different pH values (2, 7, 9, 11, 13) were evaluated on commercial whey proteins, used as a simplified protein model. The main structure and chemical modifications concerning protein aggregation, hydrolysis, insolubilization, amino acid degradation and racemization were investigated in detail. Using in vitro static models, the degree of protein hydrolysis and the released peptides were determined after the digestive process. Accumulation of molecular modifications was mostly observed after basic pH and high temperatures treatments, together with a marked decrease and modification of the digestibility profile. Instead, protein digestibility increased in neutral and acidic conditions in a temperature-dependent manner, even if some modification in the structure occurs., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The current production of meat presents many disadvantages for the environment and much research focuses on alternative protein sources. Insects are novel protein sources highly valued for their nutritional and sustainable potential. However, many aspects concerning biological and nutritional properties of the insects after digestion, in comparison with other protein sources, are still overlooked. In this work, a comparative study on three different protein sources, namely almond, lean beef and insect Alphitobius diaperinus (lesser mealworm), was performed after in vitro simulated gastrointestinal digestion. An in-depth characterization of the chemical composition of the solubilized protein and lipid fractions of the digesta was performed by applying different analytical techniques, including chromatographic methods coupled to mass spectrometry and 1 H NMR spectroscopy. Beef and insect were proven to be very similar in amino acid composition and protein solubilization after digestion, when considering the proper corrections for the chitin content. Lipid fraction from insects was solubilized during digestion as the one of almonds, but with a fastest kinetics. Thus, lesser mealworms are a good source of both lipids and highly nutritional proteins. Then, the amino acid composition of raw and digested protein fraction from the three sources was related to the PYY, ghrelin, GLP-1 and CCK release and rats' food intake. The composition of amino acids in insect digesta was found to be related to specific effects on enterohormone release, and the modulation of food intake in rats., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Black soldier fly (BSF) is being increasingly used for agro-food by-products valorisation. Adult flies, puparia, and excess of prepupae are the by-products of this process, which could be further valorised. Lactic fermentation of BSF biomasses with two different strains ( L. rhamnosus and L. plantarum ) has been used for this purpose. Deep changes in the molecular composition were observed, without significant differences related to the different strains used. The lipid and protein fractions were the most impacted. Fermentation enriched the biomass in monounsaturated and polyunsaturated fatty acids and essential amino acids, significantly improving the nutritional properties of the substrates. Although not particularly marked, a proteolytic activity of lactobacilli was observed on the BSF muscular and cuticular proteins, especially in the samples of adult flies and puparia, where fermentation resulted more effective. Conversely, there was no evidence of chitinolytic activity., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Author(s).)

The effect of thermal processing on digestibility of milk proteins should be better understood as this can greatly affect their immunoreactivity. The aim of this study was to evaluate the effects of thermal processing and lactosylation on digestibility and allergenicity, by comparing non heat-treated with industrially processed whey proteins. A semi-dynamic model was used to mimic the kinetics of digestion, and ELISA inhibition tests against human specific serum IgE were performed on the mass-spectrometry characterized products. A quicker gastric digestion of the industrially treated sample produced a lower immunogenic response in comparison with the raw sample, where intact conformational epitopes remained. In later digests, greater IgE reactivity was shown in the heat treated product, probably due to the release of linear epitopes, while at intestinal level the immunogenic response was negligible. Moreover, transepithelial transport of a reported β-lactoglobulin-derived allergen, KIDALNENVLVL, produced during digestion was assayed. It was found that the epitope-belonging peptide could be transported through the cell monolayer, both in the native and mono-lactosylated forms, with a favored passage of the native peptide.

This chapter aims to address an issue of ancient origins, but more and more topical in a globalized world in which consumers and stakeholders are increasingly aware: the authenticity of food. Foods are systems that can also be very complex, and verifying the correspondence between what is declared and the actual characteristics of the product is often a challenging issue. The complexity of the question we want to answer (is the food authentic?) means that the answer is equally articulated and makes use of many different analytical techniques. This chapter will consider the chemical analyses of foods aimed at guaranteeing their authenticity and will focus on frontier methods that have been developed in recent years to address the need to respond to ever-increasing guarantees of authenticity. Targeted and non-targeted approaches will be considered for verifying the authenticity of foods, through the study of different classes of constituents (proteins, metabolites, lipids, flavors). The numerous approaches available (proteomics, metabolomics, lipidomics) and the related analytical techniques (LC-MS, GC-MS, NMR) are first described from a more general point of view, after which their specific application for the purposes of authentication of food is addressed., (Copyright © 2022 Elsevier Inc. All rights reserved.)

In 2020, the number of deaths increased in Italy, mainly because of the COVID-19 pandemic; mortality was among the highest in Europe, with a clear heterogeneity among regions and socio-demographic strata. The present work aims to describe trends in mortality and to quantify excess mortality variability over time and in relation to demographics, pre-existent chronic conditions and care setting of the Emilia-Romagna region (Northern Italy). This is a registry-based cross-sectional study comparing the 2020 observed mortality with figures of the previous five years by age, sex, month, place of death, and chronicity. It includes 300,094 deaths in those 18 years of age and above resident in the Emilia-Romagna region. Excess deaths were higher during the first pandemic wave, particularly among men and in March. Age-adjusted risk was similar among both men and women (Mortality Rate Ratio 1.15; IC95% 1.14-1.16). It was higher among females aged 75+ years and varied between sub-periods. Excluding COVID-19 related deaths, differences in the risk of dying estimates tended to disappear. Metabolic and neuropsychiatric diseases were more prevalent among those that deceased in 2020 compared to the deaths that occurred in 2015-2019 and therefore can be confirmed as elements of increased frailty, such as being in long-term care facilities or private homes as the place of death. Understanding the impact of the pandemic on mortality considering frailties is relevant in a changing scenario.

Not all the coffee produced goes to the roasting stage, because non-compliant green coffee beans are usually discarded by roasters and the silverskin of the coffee is usually removed and discarded. In the present work, non-compliant green coffee beans and coffee silverskins were fully characterized from a chemical point of view. In addition, enzyme-assisted extraction was applied to recover a fraction rich in proteins and polyphenols, tested for antimicrobial, antityrosinase, and antioxidant activities. Non-compliant green coffee beans showed higher amounts of polyphenols, flavanols, flavonoids, and caffeine than coffee silverskins (which were richer in tannins). The enzymatic extraction of non-compliant coffee green beans produced extracts with a good protein content and with a consistent quantity of polyphenols. The extract showed antioxidant, antityrosinase, and antimicrobial activity, thus representing a promising strategy to recover defective green coffee beans. The antioxidant and antimicrobial activity of coffee silver skins is lower than that of non-compliant coffee green beans extracts, while the antityrosinase activity is comparable.

Insects are becoming increasingly relevant as protein sources in food and feed. The Black Soldier Fly (BSF) is one of the most utilized, thanks to its ability to live on many leftovers. Vegetable processing industries produce huge amounts of by-products, and it is important to efficiently rear BSF on different substrates to assure an economical advantage in bioconversion and to overcome the seasonality of some leftovers. This work evaluated how different substrates affect the protein and amino acid content of BSF. BSF prepupae reared on different substrates showed total protein content varying between 35% and 49% on dry matter. Significant lower protein contents were detected in BSF grown on fruit by-products, while higher contents were observed when autumnal leftovers were employed. BSF protein content was mainly correlated to fibre and protein content in the diet. Among amino acids, lysine, valine and leucine were most affected by the diet. Essential amino acids satisfied the Food and Agricultural Organization (FAO) requirements for human nutrition, except for lysine in few cases. BSF could be a flexible tool to bio-convert a wide range of vegetable by-products of different seasonality in a high-quality protein-rich biomass, even if significant differences in the protein fraction were observed according to the rearing substrate.

Legumes have been known for centuries for their good nutritional properties. Unfortunately, during processing, from 5 to 25% of this production is wasted, generating by-products that can still be a rich source of useful compounds, such as proteins, which can still be used in food and feed formulations. The choice of the extraction technique is important to preserve the nutritional value of proteins since drastic conditions of pH and/or temperature could damage them. In this work, two mild extraction techniques (direct assisted extraction-DAE and enzymatic assisted extraction-EAE) were applied for protein extraction from legume by-products obtained from agro-industrial processes. The quality of proteins was evaluated considering protein integrity [SDS-PAGE, degree of hydrolysis (DH), free amino acid content, racemization degree] and nutritional features [amino acid score (AAS), digestibility]. Direct assisted extraction is the technique that has best preserved protein integrity (1-5% DH and free amino acid content <1%), The digestibility of proteins extracted with EAE is higher (no protein bands detected in SDS-PAGE) than with the one of DAE extracts, making this technique particularly suitable for those food and feed formulation were a high digestibility of proteins is required., Competing Interests: MB was employed by the company Conserve Italia Soc. Coop. Agricola. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Prandi, Zurlini, Maria, Cutroneo, Di Massimo, Bondi, Brutti, Sforza and Tedeschi.)

Brine soaking is one of the most important steps in the production of Parmigiano Reggiano cheese, since it determines the amount of salt in the final product. Reduction in salt in Parmigiano Reggiano cheese might be important for improving its nutritional profile, but it could affect the manufacturing processes by altering proteolysis and consequently the product quality. In this study, for the first time, salt reduction was explored at the industrial level on real cheese samples manufactured in a local dairy. In particular, 20 wheels were produced with conventional (18 days, 10 wheels) and shorter (12 days, 10 wheels) brining steps. In every group, wheels were studied at two different ripening times, 15 and 30 months. A shorter brining time resulted in an average 12% decrease in salt content. A full characterization of free amino acids and peptides was performed by LC-MS on all samples. Free amino acids and peptides, as expected, increased with ripening, due to proteolysis, with samples having low salt content showing a slightly faster increase when compared to standard ones, hinting to a slightly accelerated proteolytic process. Nonetheless, low-salt and conventional cheeses shared similar sensory profiles at both ripening times.

The reasons behind the increasing prevalence of celiac disease (CD) worldwide are still not fully understood. This study adopted a multilevel approach (in vitro, ex vivo, in vivo) to assess the potential of gluten from different wheat varieties in triggering CD. Peptides triggering CD were identified and quantified in mixtures generated from simulated gastrointestinal digestion of wheat varieties ( n = 82). Multivariate statistics enabled the discrimination of varieties generating low impact on CD (e.g., Saragolla) and high impact (e.g., Cappelli). Enrolled subjects ( n = 46) were: 19 healthy subjects included in the control group; 27 celiac patients enrolled for the in vivo phase. Celiacs were divided into a gluten-free diet group (CD-GFD), and a GFD with Saragolla-based pasta group (CD-Sar). The diet was followed for 3 months. Data were compared between CD-Sar and CD-GFD before and after the experimental diet, demonstrating a limited ability of Saragolla to trigger immunity, although not comparable to a GFD. Ex vivo studies showed that Saragolla and Cappelli activated immune responses, although with great variability among patients. The diverse potential of durum wheat varieties in triggering CD immune response was demonstrated. Saragolla is not indicated for celiacs, yet it has a limited potential to trigger adverse immune response.

Worldwide, thousands of insect species are consumed as food or are used as feed ingredients. Hermetia illucens , 'black soldier fly', is one of them, and a large amount of puparia and dead adults flies are accumulated during rearing. These materials represent important wastes but no studies are still present in the literature regarding their functional properties and potential reuse. Lactic acid bacteria (LAB) are a heterogeneous group of bacteria contributing to various industrial applications, ranging from food fermentation, chemicals production to pharmaceuticals manufacturing. A LAB feature of industrial interest is their ability to produce antimicrobial metabolites. Considering the scientific and commercial interest in discovering novel antimicrobials, this work will be direct towards fermentation of insect-derived biomasses: puparia and adults insect at the end of life cycle. To the best of our knowledge, the in vitro antimicrobial activity of fermented insects is tested for the first time. This study aimed also to evaluate differences in the composition between fermented and unfermented insects, and to study whether the fermentation and the type of LAB used played a crucial role in modifying the composition of the substrate. Results firstly highlighted fermentability of this species of insects, showed that fermented black soldier flies puparium possess a high antimicrobial activity against tested pathogens. Moreover, result of chemical composition showed that fermented biomass had a higher percentage of fat and a more complex fatty acids profile.

Cheese microbiota contribute significantly to the final characteristics of cheeses due to the growth and interaction between cheese microorganisms during processing and ripening. For raw milk cheeses, such as Parmigiano Reggiano (PR), the microbiota derive from the raw milk itself, the dairy environment, and the starter. The process of cheese making and time of ripening shape this complex ecosystem through the selection of different species and biotypes that will drive the quality of the final product by performing functions of their metabolism such as proteolysis. The diversity in the final peptide and amino acid composition of the cheese is thus mostly linked to the diversity of this microbiota. The purpose of this study was to get more insight into the factors affecting PR cheese diversity and, more specifically, to evaluate whether the composition of the bacterial community of cheeses along with the specific peptide composition are more affected by the ripening times or by the cheese making process. To this end, the microbiota and the peptide fractions of 69 cheese samples (from curd to cheese ripened 24 months) were analyzed during 6 complete PR production cycles, which were performed in six different dairies located in the PR production area. The relation among microbial dynamics, peptide evolution, and ripening times were investigated in this unique and tightly controlled production and sampling set up. The study of microbial and peptide moieties in products from different dairies - from curd to at least 12 months, the earliest time from which the cheese can be sold, and up to a maximum of 24 months of ripening - highlighted the presence of differences between samples coming from different dairies, probably due to small differences in the cheese making process. Besides these differences, however, ripening time had by far the greatest impact on microbial dynamics and, consequently, on peptide composition., (Copyright © 2020 Bottari, Levante, Bancalari, Sforza, Bottesini, Prandi, De Filippis, Ercolini, Nocetti and Gatti.)

Insects have been proposed as new source of proteins to meet the growing demand connected to the increasing world population. In the EU the inclusion of insect proteins in feed and food is strictly regulated. Hence, analytical methods able to discriminate and identify different insect species in food and feed are a necessity. In this work, a peptidomic approach was applied to determine peptide biomarkers for two edible insect species: lesser mealworm and black soldier fly. Three species specific peptide biomarkers were identified using LC-MS/MS. The two insects were mixed with fish standard feed at different concentrations, to evaluate the feasibility of their use as markers in complex matrices. The detection of marker peptides was confirmed down to 1% insect amount. The data here reported constitutes the first proof of concept for the potential application of the peptide marker approach for the identification and quantification of insect ingredients., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Protein hydrolysates from lesser mealworm ( Alphitobius diaperinus , LM) were obtained by enzymatic hydrolysis with protease from Bacillus licheniformis . A preliminary test performed for five hours of hydrolysis generated an insect protein hydrolysate with 15% of degree of hydrolysis (DH), optimum solubility property and oil holding capacity, but emulsifying and foaming ability were completely impaired. In order to investigate the potential implication of DH on techno-functional properties, a set of protein hydrolysates with a different DH was obtained by sub-sampling at different time points during three hours of enzymatic hydrolysis process. An increase in DH% had positive effects on the solubility property and oil holding ability, while a reduced emulsifying ability was observed up to five hours of hydrolysis. These results demonstrated that the enzymatic hydrolysis, if performed under controlled conditions and not for a long period, represents a valid method to extract high quality protein from insects with tailored techno-functionality, in order to produce tailored ingredients for feed and food purpose.

The high temperatures used in the production of milk may induce modifications in proteins structure. Due to occurrence of the Maillard reaction, lactose binds lysine residues in proteins, affecting the nutritional value. Milk is also an important source of allergenic proteins (i.e., caseins, β-lactoglobulin and α-lactalbumin). Thus, this modification may also affect the allergenicity of these proteins. Focusing on milk whey proteins, a screening on different Ultra High Temperatures (UHT) and pasteurized milk samples was performed to identify lactosylation sites, in particular in protein known epitopes, and to verify the correlation between lactosylation and the harshness of the treatment. Whey proteins were extracted from milk samples after caseins precipitations at pH 4.6 and, after chymotryptic and tryptic in solution digestion, peptides were analysed by UPLC-MS and LTQ-Orbitrap. Results show the presence of lactosylated lysine residues in several known epitopes. Then, a β-lactoglobulin epitope was selected and synthesized by solid phase synthesis followed by in solution lactosylation, obtaining high reaction yields and purities. The synthesis of lactosylated allergenic epitopes, described here for the first time, is a useful tool for further studies on the technological impacts on food allergenicity.

Parmigiano-Reggiano (PR) is a worldwide known Italian, long ripened, hard cheese. Its inclusion in the list of cheeses bearing the protected designation of origin (PDO, EU regulation 510/2006) poses restrictions to its geographic area of production and its technological characteristics. To innovate the Parmigiano-Reggiano (PR) cheese manufacturing chain from the health and nutritional point of view, the output of defatted PR is addressed. Two defatting procedures (Soxhlet, and supercritical CO 2 extraction) were tested, and the obtained products were compared in the composition of their nitrogen fraction, responsible for their nutritional, organoleptic, and bioactive functions. Free amino acids were quantified, and other nitrogen compounds (peptides, proteins, and non-proteolytic aminoacyl derivatives) were identified in the extracts and the mixtures obtained after simulated gastrointestinal digestion. Moreover, antioxidant and angiotensin converting enzyme (ACE) inhibition capacities of the digests were tested. Results obtained from the molecular and biofunctional characterization of the nitrogen fraction, show that both the defatted products keep the same nutritional properties of the whole cheese.

Enzymatic improvers are enzymes obtained from microbial or fungal cultures, added as technical adjuvants to flour, with the aim of improving the dough characteristics in bakery products. They are used in a low ppm range and, being technical adjuvants, can go undeclared on the label. Many types of enzymatic improvers are present on the market, such as amylases, lipases, proteases, xylanases, glucose oxidases, and others, each with a different function. Analytical methods capable of detecting these enzymes are needed, particularly for bakery companies, in order to monitor the quality of raw materials and to detect any undeclared presence. In the present work, specific peptide markers, obtained by enzymatic digestion, have been used to detect the presence of enzymatic improvers by LC-MS/MS techniques. Promising results were obtained for some enzymes acting on the carbohydrate fraction (glucoamylase, glucose oxidase, xylanase) in which amounts as low as 20 ppm could be identified in blind flour samples. For lipases and proteases the method proved to be very effective in terms of specific identification, even if less sensitive.

Since 2018, insects have belonged the category of Novel Foods and the presence of allergens represents one of the main hazards connected to their consumption, also due to the potential cross-reactivity with Arthropoda pan-allergens. In the present work, the allergenicity assessment of black soldier fly and lesser mealworm was performed with a shotgun bottom-up proteomic approach combined with in-silico assessment, followed by IgG- and IgE-immunoblotting experiments. The peptides identified, filtered for their abundance and robustness, belonged mainly to muscle proteins, which represented the most abundant protein group. The relevant potential allergens were in-silico identified by sequence similarity to known allergens, and among them tropomyosin resulted the most abundant insect allergen. IgG-immunoblotting analysis with anti-Tropomyosin I antibodies and IgE-immunoblotting assay with serum from patient allergic to crustacean tropomyosin were performed in order to assess the immunoreactivity in both insects. The immunoassays were carried out also on protein hydrolysates extracted by treating insects with Protease from Bacillus licheniformis (1%, 60 °C, pH 7.5). While IgG-immunoblotting demonstrated the loss of immunoreactivity for both hydrolysates, IgE-immunoblotting showed a partial immunoreactivity preservation, also after hydrolysis, in the case of black soldier fly hydrolysate, and a total loss of immunoreactivity for lesser mealworm hydrolysate.

Insects are considered a suitable alternative feed for livestock production and their use is nowadays regulated in the European Union by the European Commission Regulation No. 893/2017. Insects have the ability to grow on a different spectrum of substrates, which could be naturally contaminated by mycotoxins. In the present work, the mycotoxin uptake and/or excretion in two different insect species, Alphitobius diaperinus (Lesser Mealworm, LM) and Hermetia illucens (Black Soldier Fly, BSF), grown on naturally contaminated substrates, was evaluated. Among all the substrates of growth tested, the Fusarium toxins deoxynivalenol (DON), fumonisin 1 and 2 (FB1 and FB2) and zearalenone (ZEN) were found in those based on wheat and/or corn. No mycotoxins were detected in BSF larvae, while quantifiable amount of DON and FB1 were found in LM larvae, although in lower concentration than those detected in the growing substrates and in the residual fractions. Mass balance calculations indicated that BSF and LM metabolized mycotoxins in forms not yet known, accumulating them in their body or excreting in the faeces. Further studies are required in this direction due to the future employment of insects as feedstuff.

Resistance of gluten to gastrointestinal digestion is involved in immune-mediated adverse reactions to wheat, since several peptides produced by the incomplete digestion are able to trigger, in predisposed individuals, the immune response responsible, for instance, of celiac disease (CD) and other adverse reactions. Even if several peptides have been identified, an exhaustive description of the peptidome generated by wheat digestion is lacking. To this end, in the present work, durum wheat proteins were fractionated, digested, and then subjected to various proteomic techniques, including single stage and multiple stage mass spectrometry (MS) (SDS-PAGE, UPLC/ESI-MS, UPLC/ESI-MS/MS, and LTQ-Orbitrap). Based on SDS-PAGE, although proteins were severely degraded after in vitro gastrointestinal digestion, some differences were observed among protein profile of the different digests. Through untargeted UPLC techniques, 227 peptide sequences were identified, with only few sequences shared by the different digests. In particular, 9 gluten peptides involved in CD were identified. Based on target proteomic, the quantification of these peptides revealed significant (p ≤ 0.05) differences among the different extracts. Taken together, all the proteomic tools confirmed that gluten digestion is closely related to the matrix regardless of wheat genotype.

The nitrogen fraction of 39 food waste streams was characterized by Kjeldahl analysis, amino acid analysis, protein analysis and racemization degree, for assessing their potential for further valorization. For every waste streams the specific nitrogen-to-protein conversion factor was calculated, allowing to assess the accurate protein content. The results indicated which streams are most rich in relevant proteins (all wastes of dairy origin, beer yeast, malted barley germs, brewing cake, rapeseed press cake, sea buckthorn spent pulp, leek leaves, parsley waste, pumpkin kernel cake, and mushroom waste), which ones have valuable proteins, but in too little amount, and also which ones are rich in proteins, but of low nutritional value. Specific data also indicated, for every waste stream, its possible use for supplementing specific amino acids. To date, this represents the most complete characterization with homogeneous methodologies of the nitrogen fraction in food waste streams ever reported in the literature and outlines in unprecedented molecular details the potentialities and the limitations of many waste streams to be used as source of proteins., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Old wheat genotypes are perceived by consumers as healthier than modern ones. The release of gluten peptides with in vitro digestion and the content of potentially prebiotic carbohydrates (i.e. resistant fraction of starch and cell-wall associated dietary fiber) were evaluated in tetraploid wheats, namely 9 old and 3 modern Triticum turgidum ssp. genotypes. Simulated digestion of wholemeal flours yielded 152 major peptides, 59 of which were attributed a sequence. Principal component analysis revealed that peptide profiles were variable in old genotypes, unlike in modern ones. Digestion of old genotypes generally yielded peptides in greater concentration. In particular, 5 peptides of γ-gliadin, known to trigger the adaptive immune reaction, and two peptides of α-gliadin, known to be toxic to celiac patients, were particularly abundant in some old varieties. Resistant starch (RS) was negligible in modern genotypes (<0.6%), but it was remarkably abundant in some old varieties, reaching the highest value in Dauno III (8.5%, P < 0.05). Dauno III also presented the highest amount of soluble fiber (4.2%, P < 0.05). Pasta was made with an old and a modern genotype (Dauno III and PR22D89, respectively) with opposite RS content. Pasta making and cooking affected starch digestibility, overtaking differences between genotypes and yielding the same amount of RS for both the varieties (approx. 1.7%). The data herein presented suggest that the wholemeal flours of old tetraploid wheat genotypes could not boast particular claims associated to a lower exposure to gluten peptides and, if cooked, to a prebiotic potential., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

In this study we investigated the oligopeptide pattern in fermented cocoa beans and derived products after simulated gastrointestinal digestion. Peptides in digested cocoa samples were identified based on the mass fragmentation and on the software analysis of vicilin and 21 KDa cocoa seed protein sequences, the most abundant cocoa proteins. Quantification was carried out by liquid chromatography/electrospray ionisation mass spectrometry (LC/ESI-MS) using an internal standard. Sixty five peptides were identified in the digested samples, including three pyroglutamyl derivatives. The in vitro angiotensin-converting enzyme (ACE)-inhibitory activity of cocoa digests were tested, demonstrating a high inhibition activity, especially for digestates of cocoa beans. The peptides identified were screened for their potential ACE inhibitory activity through an in silico approach, and about 20 di-, three- and tetra-peptides actually present in our samples were predicted as active. Two of the potentially active peptides were chemically synthesized and then assessed for their inhibitory activity by using the ACE in vitro assay. These peptides demonstrated an ACE inhibitory activity, however, that was too weak to explain alone the high activity of cocoa digestates, suggesting a synergic effect of all cocoa peptides. As a whole, results showed that an average chocolate portion (30 g) ensures an amount of peptides after digestion that, assuming complete absorption, could reach almost a complete inhibition of ACE.

Durum wheat ( Triticum turgidum L. subsp. durum (Desf.) Husn) is a major food source in Mediterranean countries since it is utilized for the production of pasta, leavened and unleavened breads, couscous, and other traditional foods. The technological and nutritional properties of durum wheat semolina depend mainly on the type of gluten proteins and on their amount, which is a genotype- and environment-dependent trait. Gluten proteins are also responsible for celiac disease (CD), an autoimmune enteropathy with a prevalence of about 0.7-2% in the human population. At this purpose, two Italian durum wheat cultivars, Saragolla and Cappelli, currently used for monovarietal pasta, were chosen to compare (i) the reserve and embryo proteome, (ii) the free and bound phenolics, antioxidant activity, and amino acid composition, and (iii) the content of immunogenic peptides produced after a simulated gastrointestinal digestion. The results obtained from 2 years of field cultivation on average showed a higher amount of gluten proteins, amino acids, and immunogenic peptides in Cappelli. Saragolla showed a higher abundance in bound phenolics, antioxidant enzymes, and stress response proteins in line with its higher antioxidant activity. However, the impact of the year of cultivation, largely depending on varying rainfall regimes through the wheat growth cycle, was significant for most of the parameters investigated. Differences in technological and nutritional characteristics observed between the two cultivars are discussed in relation to the influence of genetic and environmental factors.

Black soldier fly (BSF, Hermetia illucens) represents a valuable source of biomolecules and it also constitutes an economic way to valorise residual biomasses. BSF prepupae contain high amounts of lipids (37% DM basis). The present investigation aimed at studying the composition of BSF lipids and the effect of killing/storage on their quality. The main fatty acid was lauric acid, sterols were represented primarily by beta-sitosterol and campesterol. Global fatty acid and sterol profiles, determined by GC-MS, were only slightly affected by the killing procedure, while lipid classes distribution, determined by 1 H NMR, strongly changed. Prepupae killed by freezing showed a drastic reduction of acylglycerols during storage and a relevant release of free fatty acids, likely due to activation of lipases. On the contrary, prepupae killed by blanching have a stable lipid fraction constituted mainly by triacylglycerols. Therefore, killing procedure strongly influences BSF oil composition and the potential applications., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Insects are being explored as novel protein sources in order to overcome the future food demands connected to world growing population. Insects for food/feed uses are currently slowly killed through freezing by most insect rearing companies, and typically, enzymatic browning takes place in the insect proteins fractions. However, very little is known about the influence of these enzymatic reactions on the protein physical, chemical, nutritional and technological properties. In this work a metabolomics and proteomic study was conducted on Black Soldier Fly (Hermetia illucens) prepupae, killed by two different methods: freezing (commonly used), and blanching (with the aim to inhibit the enzymatic activities). Proton nuclear magnetic resonance ( 1 H NMR) metabolomics demonstrated that slow killing method by freezing, compared with blanching, elicits the activation of several enzymatic pathways, among them melanisation with tyrosine consumption, energetic metabolism and lipolysis. These metabolic changes have an impact also on protein nutritional quality, with a loss of cysteine and lysine, likely involved in the process of melanisation and enzymatic browning. A strong effect was also observed on protein extractability: proteins from prepupae killed by blanching were found to be more extractable in milder conditions by chemical methods, and more prone to enzymatic digestion (97% of proteins released in solution upon proteolysis) than proteins from prepupae killed by freezing. All these data indicate that killing by blanching inhibits the browning reaction and other enzymatic changes occurring during slow killing by freezing, increasing the extractability of proteins in aqueous solutions, avoiding essential amino acid loss, and improving enzymatic digestibility., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Proteolysis is the most important event occurring during maturation of dry-cured hams: it strongly influences the flavour and the texture of the aged ham by the accumulation of peptides and free amino acids released by protein hydrolysis. Apart from compounds of proteolytic origin, it has been demonstrated that also non-proteolytic amino acyl derivatives (γ-glutamyl amino acids, pyroglutamyl-amino acids and lactoyl-amino acids) may accumulate during ripening of cheese, and they can be also found in fermented soy sauce, where they contribute to the umami taste of the products. Using a semi-quantitative analysis, in this paper we report the occurrence of significant amounts of γ-glutamyl amino acids and, for the first time, pyroglutamyl-amino acids and lactoyl-amino acids, in aged ham. The amino acid counterparts were mainly found to be hydrophobic amino acids. The amount of these compounds was found to increase with time, because they are not degraded by proteolytic activity. They were also found to be stable to simulated gastrointestinal digestion. Angiotensin Converting Enzyme inhibitory activity was also tested, but they were not found to be characterized by significant ACE-inhibitory activity., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The aim of this research was to valorize onion skins, an under-utilized agricultural by-product, into pectic oligosaccharides (POS), compounds with potential health benefits. To achieve high hydrolysis performance with the multi-activity enzyme Viscozyme L, an innovative approach was investigated based on a cross-flow continuous membrane enzyme bioreactor (EMR). The influence of the various process conditions (residence time, enzyme concentration, substrate concentration) was investigated on productivity and yield. The composition of the POS mixtures in terms of mono- and oligosaccharides was assessed at the molecular level. At optimized conditions, a stable POS production with 22.0g/L/h volumetric productivity and 4.5g/g POS/monosaccharides was achieved. Compared to previous results obtained in batch for the enzyme Viscozyme L, EMR provided a 3-5× higher volumetric productivity for the smallest POS. Moreover, it gave competitive results even when compared to batch production with a pure endo-galacturonase enzyme, demonstrating its feasibility for efficient POS production., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Sugar beet pulp pectin is an attractive source for the production of pectic oligosaccharides, an emerging class of potential prebiotics. The main aim of the present work was to investigate a new process allowing to produce pectic oligosaccharides in a continuous way by means of a cross flow enzyme membrane reactor while using a low-cost crude enzyme mixture (viscozyme). Preliminary experiments in batch and semi-continuous setups allowed to identify suitable enzyme concentrations and assessing filtration suitability. Then, in continuous experiments in the enzyme membrane reactor, residence time and substrate loading were further optimized. The composition of the obtained oligosaccharide mixtures was assessed at the molecular level for the most promising conditions and was shown to be dominated by condition-specific arabinans, rhamnogalacturonans, and galacturonans. A continuous and stable production was performed for 28.5 h at the optimized conditions, obtaining an average pectic oligosaccharide yield of 82.9 ± 9.9% (w/w), a volumetric productivity of 17.5 ± 2.1 g/L/h, and a specific productivity of 8.0 ± 1.0 g/g E/h. This work demonstrated for the first time the continuous and stable production of oligosaccharide mixtures from sugar beet pulp using enzyme membrane reactor technology in a setup suitable for upscaling.

Germination is already a well-accepted process by consumers with many products made from sprouted seeds or containing limited amounts of flour form sprouted grains. The present work aimed assessing the usefulness of germination in reducing gluten peptides associated with celiac disease, at the same time evaluating some technological features of the obtained germinated wheat. In the first part of the work, celiac disease (CD)-triggering peptides were tracked as a function of germination kinetics (from day 1 to day 6). Using simulated gastrointestinal digestion and liquid chromatography coupled to mass spectrometry, ten celiac disease triggering peptides were identified: seven peptides presumably involved in the adaptive immune response (TI) and three peptides mainly involved in the innate immune response (TT). All the identified peptides belonged to gliadins. TI track pattern showed three phases: the first two days displayed a significant degradation, a stability phase was observed from day 3 to day 5, and finally a drastic reduction occurred on the 6th day. For TT peptides, important degradation was exclusively observed at the 6th day. In the second part, some techno-functional features of germinated whole wheat flour were assessed to estimate its potential as an alternative to conventional flour. Functionality comparison of the non-germinated versus germinated flours revealed that germination significantly influenced solvents retention capacities as well as swelling and solubility. Thus, with a reduced amount of celiac disease triggering peptides, but also with different technological behavior compared to traditional wheat flour., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Celiac disease (CD) is an immune-mediated enteropathy triggered by ingested gluten in genetically susceptible individuals and sustained by both adaptive and innate immune responses. Recent studies in murine macrophages demonstrated that the activation of arginase (ARG) metabolic pathway by gluten peptides contributes to the modulation of intestinal permeability in vitro. Here we characterize the effects of gluten on arginine metabolism and cell polarization in human monocytes from both healthy and CD subjects; both a simplified enzymatic digestion of gliadin and a physiological digestion of whole wheat have been tested. Results indicate that gluten digests induce the onset of an M2-like phenotype in activated macrophages; more precisely, both isoforms of arginase, ARG1 and ARG2, are induced likely due to the inhibition of mTOR and the consequent induction of C/EBPβ transcription factor. These effects are independent from the origin of gluten as well as from the digestive protocol employed; moreover, no statistical difference can be evidenced between healthy and CD patients, excluding a diverse predisposition of CD monocytes to gluten-triggered polarization with respect to healthy immune cells. Overall, the present findings sustain a role for arginase pathway in the immune response elicited by human monocytes toward ingested gluten that, hence, deserves particular attention when addressing the pathogenesis of CD., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The protein content and allergen composition was studied of cashews from 8 different origins (Benin, Brazil, Ghana, India, Ivory Coast, Mozambique, Tanzania, Vietnam), subjected to different in-shell heat treatments (steamed, fried, drum-roasted). On 2D electrophoresis, 9 isoforms of Ana o 1, 29 isoforms of Ana o 2 (11 of the acidic subunit, 18 of the basic subunit), and 8 isoforms of the large subunit of Ana o 3 were tentatively identified. Based on 1D and 2D electrophoresis, no difference in allergen content (Ana o 1, 2, 3) was detected between the cashews of different origins (P > 0.5), some small but significant differences were detected in allergen solubility between differently heated cashews. No major differences in N- and C-terminal microheterogeneity of Ana o 3 were detected between cashews of different origins. Between the different heat treatments, no difference was detected in glycation, pepsin digestibility, or IgE binding of the cashew proteins., (© 2018 Institute of Food Technologists®.)

Black soldier fly (BSF, Hermetia illucens) constitutes an economic way to convert residual biomasses into a valuable source of biomolecules, such as proteins, lipids and chitin. The present investigation was undertaken to evaluate the feasibility of applying different extraction protocols, either chemical extractions or enzymatic assisted extraction, to recover pure fat, protein and chitin fractions. First, exact proximate composition, total amino acids, fatty acids profile, and N-acetylglucosamine content of the prepupae samples were determined. BSF prepupae biomass contained, expressed on dry weight, 32% proteins, 37% lipids, 19% minerals, 9% chitin. The lipid fraction was easily recovered by organic solvents, while the most challenging issue was the separation of protein from chitin. The best separation was obtained by alkali extraction of proteins (96% of protein recovered) albeit with loss in their integrity as indicated by the measurement of the degree of hydrolysis with the o-phthaldialdehyde method. To avoid protein damage in alkali media, a stepwise protein extraction adopting milder conditions was also explored based on Osborne fractionation method, allowing the recovery of >85% of BSF high purity and high quality proteins, and the obtainment of chitin-enriched fraction as well. The possibility of using an enzymatic assisted extraction of proteins was also explored, obtaining a maximum nitrogen solubilisation in the best case (with Bacillus licheniformis protease) of about 60%. In this latter case, the chitin fraction obtained also had a significant residual protein content., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Pectin oligosaccharides (POS) have been indicated as a new class of potential prebiotic compounds, which can be produced from pectin-rich food byproducts. In the present study, different technological means of POS production were explored to produce tailor-made POS mixtures starting from sugar beet pulp. The overall POS production process consisted of two steps: the extraction of pectin and the hydrolysis of pectin to tailored POS by combined hydrolysis/fractionation approaches. Different extraction as well as hydrolysis and fractionation methodologies were applied. The obtained POS were characterized for their total galacturonic acid content and, at a deeper level, using a HILIC-ESI/MS methodology, for the POS structure and composition. The composition of POS fractions was studied as a function of the technology used to obtain them. Finally, the potential prebiotic properties of the POS mixtures obtained were thoroughly explored by several in vitro experiments aimed at detecting lactic acid bacteria (LAB) stimulation by POS fractions. Several fractions were very efficient in stimulation, in a species-dependent manner. The overall best fractions were in general those rich in arabinans having a low degree of polymerization, obtained from the enzymatic extraction of biomass and subsequent fractionation with low-medium molecular weight cut-off. Quite interestingly, no POS fraction was able to stimulate pathogenic E. coli strains. The data reported here clearly indicate the possibility to obtain diverse fractions with different prebiotic properties starting from the same biomass, and outline clear potential for POS obtained from sugar beet pulp with the appropriate technology to act as prebiotic compounds.

For centuries, ancient grains fed populations, but due to their low yield, they were abandoned and replaced by high-yielding species. However, currently, there is a renewed interest in ancient wheat and pseudocereal grains from consumers, farmers, and manufacturers. Ancient wheat such as einkorn, emmer, spelt, and Kamut®, are being reintegrated because of their low fertilizer input, high adaptability and important genetic diversity. New trends in pseudocereal products are also emerging, and they are mostly appreciated for their nutritional outcomes, particularly by the gluten-free market. Toward healthier lifestyle, ancient grains-based foodstuffs are a growing business and their industrialization is taking 2 pathways, either as a raw ingredient or a functional ingredient. This paper deals with these grain characteristics by focusing on the compositional profile and the technological potential., (© 2017 Institute of Food Technologists®.)

In this work, the aim is to study the effectiveness of germination on wheat protein degradation, with a specific focus on proteins involved in adverse reactions to wheat. The effects of 8 days of germination at 25 °C on the chemical composition and the protein profile were determined. Germination did not have a significant effect on starch, protein, lipid, and ash contents. General protein profile, as indicated by SDS-PAGE analysis, revealed that germination induced a relevant degradation in protein fraction. After in vitro gastrointestinal digestion, gluten peptides involved in celiac disease (CD) were identified and quantified using UPLC/ESI-MS technique. Also, CM3 protein, involved in baker's asthma and intestinal inflammation, was quantified by measuring a marker peptide. Statistical analysis underlined that germination and genotype had significant impact on the amount of both components. Regarding gluten peptides related to CD, germination enabled an average reduction of 47% in peptides eliciting adaptive immune response and 46% in peptides eliciting innate immune response. CM3 protein showed also a high average reduction (56%). Thus, this study suggests that germination might be a good bioalternative to provide a low "impact" raw ingredient for special wheat-based foodstuffs.

Introduction: Phlorotannins are complex mixtures of phloroglucinol oligomers connected via C-C (fucols) or C-O-C (phlorethols) linkages. Their uniformity in subunits and large molecular weight hamper their structural analysis. Despite its commercial relevance for alginate extraction, phlorotannins in Laminaria digitata have not been studied., Objective: To obtain quantitative and structural information on phlorotannins in a methanolic extract from L. digitata., Methodology: The combined use of 13 C and 1 H NMR spectroscopy allowed characterisation of linkage types and extract purity. The purity determined was used to calibrate the responses obtained with the colorimetric 2,4-dimethoxybenzaldehyde (DMBA) and Folin-Ciocalteu (FC) assays. Using NP-flash chromatography, phlorotannin fractions separated on oligomer size were obtained and enabled structural and molecular weight characterisation using ESI-MS and MALDI-TOF-MS., Results: The fucol-to-phlorethol linkage ratio was 1:26 and the extract was 60.1% pure, determined by NMR spectroscopy. For DMBA, the response of the extract was 12 times lower than that of phloroglucinol, whereas there was no difference for FC. By accounting for differences in response, the colorimetric assays were applicable for quantification using phloroglucinol as a standard. The phlorotannin content was around 4.5% DM. Fucol- and phlorethol-linkage types were annotated based on characteristic MS n fragmentations. Structural isomers of phlorotannins up to a degree of polymerisation of 18 (DP18) were annotated and identification of several isomers hinted at branched phloroglucinol oligomers. With MALDI-TOF-MS phlorotannins up to DP27 were annotated., Conclusion: By combining several analytical techniques, phlorotannins in L. digitata were quantified and characterised with respect to fucol-to-phlorethol linkage ratio, molecular weight (distribution), and occurrence of structural isomers. Copyright © 2017 John Wiley & Sons, Ltd., (Copyright © 2017 John Wiley & Sons, Ltd.)

Pectic oligosaccharides (POS) have been indicated as novel candidate prebiotics. Traditionally, POS are produced from pectin-rich by-products using a two-step process involving extraction of the pectin, followed by its hydrolysis into POS. A one-step approach, in which the POS is directly produced from the raw material, might provide a more efficient alternative. Thus, the main aim of this paper was to investigate a one-step enzymatic hydrolysis approach to directly produce POS from sugar beet pulp (SBP). The POS yield was investigated as a function of the process parameters, as well as raw material characteristics. A statistically-based response surface methodology, using a central composite design was applied, to investigate the individual as well as the combined influences of the diverse parameters. The model was confirmed by a validation experiment, carried out at 135 g/l substrate concentration, 0.75 FPU/g SBP enzyme concentration, 0.8 mm particle size and 3 h hydrolysis time. Under these conditions, a POS-rich hydrolysate was obtained, containing rhamnose, arabinose, galactose, xylose and galacturonic acid, at 0.9, 15.2, 5.1, 1.4, and 13.2 g/l, respectively, enzymes were added each at 20 FPU/g dry matter (DM).

Because of the continuous increase in the prevalence of gluten-related disorders, selection of wheat with a low content of immunogenic gluten epitopes could be an innovative alternative for prevention. In this review, the focus is on literature data concerning the deallergenization tools of wheat, which are mainly related to breeding approaches (classic and advanced) and processing operations (germination and fermentation). Until now, no safe wheat genotype has been identified, whereas decreasing wheat allergenicity is possible. On the other hand, the decrease of gluten or some of its epitopes can strongly affect technological properties. Thus, obtaining celiac-safe gluten without affecting the technological properties of wheat could be considered as a new challenge that scientists will be facing. Celiac-safe wheat-based product development could be a great revolution in the market of foods for special medical purposes. The present paper is aiming to: (a) review the strategies and the approaches used, or that can be used, for developing low allergenic wheat: their utilities and limits were also discussed and (b) screen the impact of gluten reduction or removal on the quality of wheat end-use products., (© 2017 Institute of Food Technologists®.)