Glucose/Ribitol Dehydrogenase and 16.9 kDa Class I Heat Shock Protein 1 as Novel Wheat Allergens in Baker's Respiratory Allergy.

Wheat allergens are responsible for symptoms in 60-70% of bakers with work-related allergy, and knowledge, at the molecular level, of this disorder is progressively accumulating. The aim of the present study is to investigate the panel of wheat IgE positivity in allergic Italian bakers, evaluating a possible contribution of novel wheat allergens included in the water/salt soluble fraction. The water/salt-soluble wheat flour proteins from the Italian wheat cultivar Bolero were separated by using 1-DE and 2-DE gel electrophoresis. IgE-binding proteins were detected using the pooled sera of 26 wheat allergic bakers by immunoblotting and directly recognized in Coomassie stained gel. After a preparative electrophoretic step, two enriched fractions were furtherly separated in 2-DE allowing for detection, by Coomassie, of three different proteins in the range of 21-27 kDa that were recognized by the pooled baker's IgE. Recovered spots were analyzed by nanoHPLC Chip tandem mass spectrometry (MS/MS). The immunodetected spots in 2D were subjected to mass spectrometry (MS) analysis identifying two new allergenic proteins: a glucose/ribitol dehydrogenase and a 16.9 kDa class I heat shock protein 1. Mass spectrometer testing of flour proteins of the wheat cultivars utilized by allergic bakers improves the identification of until now unknown occupational wheat allergens.

Breadstick fortification with red grape pomace: effect on nutritional, technological and sensory properties.

BACKGROUND: Grape pomace (GP), a wine-making by-product rich in dietary fiber (DF) and total phenolic compounds (TPC), is a potential functional ingredient in the fortification of baked goods. RESULTS: In the present study, fortified breadsticks samples were obtained by replacing wheat flour with 0, 5 and 10 g 100 g-1 of powdered GP (GPP). The GPP inclusion affected the rheological properties of the doughs by increasing the water absorption and tenacity (P) at the same time as reducing the extensibility (L), with a significant increase in the P/L value and a decrease in the swelling index (G) value and deformation energy (W). Textural characteristics of breadsticks were influenced by the GPP addition, showing a reduction in hardness and fracturability as the amount of GPP increased in the recipe. The GPP fortified breadsticks exhibited decreased pH, volume and specific volume values compared to the control. The TPC and the antioxidant capacity increased in GPP fortified breadsticks, whereas the increased amount of DF allowed the products to benefit from the claim 'high fiber content' at the highest level of GPP inclusion. The sensory evaluation revealed that GPP addition increased wine odor, acidity, bitterness, astringency and hardness, and decreased the regularity of alveolation and friability. Finally, the GPP fortified products achieved good sensorial acceptability. CONCLUSION: GPP improved the nutritional values of fortified breadsticks and changed the rheology of dough and breadsticks' technological properties without affecting sensory acceptability. © 2021 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Technological, nutritional, and sensory properties of durum wheat fresh pasta fortified with Moringa oleifera L. leaf powder.

BACKGROUND: Pasta is a staple food that is consumed worldwide and is an excellent product for the addition of ingredients rich in bioactive compounds. The fortification of pasta with such compounds could represent a healthy choice for consumers. RESULTS: In this study, fresh pasta was formulated by replacing durum wheat semolina with 0, 5, 10, and 15 g 100 g-1 of dried Moringa oleifera leaf powder (MOLP), rich in fibers, minerals, and antioxidant compounds. Increasing levels of MOLP influenced the technological and nutritional properties of wheat-based fresh pasta. Moringa oleifera reduced the optimum cooking time, the swelling index and firmness, while increasing the cooking loss and adhesiveness. From a nutritional viewpoint, the inclusion of MOLP enhanced the phenol content, the antioxidant activity, and the mineral content of fresh pasta. The products obtained had good sensorial acceptability and can make several nutritional claims due to MOLP richness minerals. CONCLUSIONS: The fortification of fresh pasta with MOLP could represent a valuable strategy to increase the nutritional value of the product, preserving pasta technological properties without affecting sensory acceptability. © 2020 Society of Chemical Industry.

Exploration of grape pomace peels and amaranth flours as functional ingredients in the elaboration of breads: phenolic composition, bioaccessibility, and antioxidant activity.

This study evaluated the incorporation of two ingredients as a source of bioactive compounds: amaranth flour (AF) and grape pomace peels flour (GP) to improve the nutritional qualities and functional properties of a wheat bread, emphasising the revalorisation of agricultural residues from grape winemaking as an ethical and economically viable source of bioactive compounds. Specifically, wheat flour (WF) substitutions were carried out for the individual ingredients, replacing 20% WF (A20 bread) or 5% GP (GP5 bread) and a mixture of both ingredients 20% WF and 5% GP (A20GP5 bread), and the antioxidant potential of the breads was analysed. The effect of simulated gastrointestinal digestion (SGID) on the phenolic profile and antioxidant activity of the fortified breads was also investigated. The substitution of WF by AF or GP introduced several phenolic compounds, digestion increased the bioaccessibility of phenolic compounds and reshaped their phenolic composition profiles. The combined presence of AF and GP in the breads modified the phenolic compounds composition and improved their antioxidant activity after SGID. Interactions between the phenolic compounds and other AF components (possibly proteins) were observed, which could protect the phenols from degradation during SGID, allowing them to be released after SGID.

The α-Amylase and α-Glucosidase Inhibition Capacity of Grape Pomace: A Review.

The concept of functional foods is gaining more importance due to its role in maintaining a healthy status and preventing some metabolic diseases. The control of diabetes, in particular type-2 (T2DM), could be considered a big challenge since it involves other factors such as eating habits. From the pharmacological point of view, inhibiting digestive enzymes, such as α-amylase and α-glucosidase, is one of the mechanisms mainly used by synthetic drugs to control this disease; however, several side effects are described. For that reason, using bioactive compounds may appear as an alternative without presenting the complications synthetic drugs available on the market have. The winemaking industry generates tons of waste annually, and grape pomace (GP) is the most important. GP is recognized for its nutritional value and as a source of bioactive compounds that are helpful for human health. This review highlights the importance of GP as a possible source of α-amylase and α-glucosidase inhibitors. Also, it is emphasized the components involved in this bioactivity and the possible interactions among them. Especially, some phenolic compounds and fiber of GP are the main ones responsible for interfering with the human digestive enzymes. Preliminary studies in vitro confirmed this bioactivity; however, further information is required to allow the specific use of GP as a functional ingredient inside the market of products recommended for people with diabetes.

Durum Wheat Fresh Pasta Fortification with Trub, a Beer Industry By-Product.

Trub is a brewing by-product rich in proteins and fibers. We used trub, after a debittering step, at 5, 10, and 15 g/100 g (PT5, PT10, and PT15, respectively) to fortify durum wheat fresh pasta. Technological and physical-chemical properties, in vitro digestibility, and sensorial characteristics of fortified pasta were determined. The technological aspects of the products were peculiar, suggesting the existence of complex interactions between the gluten network and starch with debittered trub powder. The fortified pasta samples showed a lower glucose release than the control at the end of in vitro starch hydrolysis. Furthermore, in vitro protein digestion rose only in PT15. PT5 and PT10 samples overcame the sensory acceptability threshold of 5, while PT15 showed the lowest acceptability. Debittered trub represents a suitable ingredient in fortified fresh pasta formulation with an up to 10% substitution level without compromising the quality and sensory characteristics of the final product.

Study of the phenolic profile of a grape pomace powder and its impact on delaying corn oil oxidation.

The grape pomace, the main by-product from the winemaking industry, contains many bioactive substances that must be valorized. The aim of this study was to assess the total phenolic content (TPC), phenolics profile by using HPLC and the antioxidant activity (AOA). The results showed a TPC of 38.86 ± 5.22 g gallic acid equivalent (GAE)/kg while an AOA of 247.84 ± 18.65 µmol Trolox equivalent (TE)/g. Epicatechins were the most representative phenolic compound, according to the HPLC analysis. Then, the grape pomace powder (GPP) was tested in the Rancimat equipment as a natural antioxidant for delaying the corn oil oxidation. Results showed statistically significant differences between the corn oil treated with GP and the control, so the GPP could be a promising natural antioxidant to tackle the oxidation vulnerability of corn oil.

Predicted Shelf-Life, Thermodynamic Study and Antioxidant Capacity of Breadsticks Fortified with Grape Pomace Powders.

Grape pomace (GP), is the main winemaking by-product and could represent a valuable functional food ingredient being a source of bioactive compounds, like polyphenols. Polyphenols prevent many non-communicable diseases and could contrast the oxidation reaction in foods. However, the high content in polyunsaturated fatty acid, the described pro-oxidant potential action of some polyphenols and the complex interactions with other components of matrices during food processing must be considered. Indeed, all these factors could promote oxidative reactions and require focused and specific assay. The aims of this study were to evaluate the effects of GP powder (GPP) addition (at 0%, 5% and 10% concentrations) in breadsticks formulations both on the antioxidant activity at room temperature during storage and on the shelf-life by the OXITEST predictive approach. GPP fortification increased the total polyphenols content and the antioxidant activities of breadsticks. FRAP reduced during the first two days of storage at room temperature, TPC increased during the 75 days, while ABTS showed a slight progressive decrease. However, GP negatively influenced OXITEST estimated shelf-life of breadsticks, incrementing the oxidation rate. In conclusion, even if GP fortification of breadsticks could improve the nutritional value of the products, the increased commercial perishability represents a drawback that must be considered.

Wheat Bread Fortification by Grape Pomace Powder: Nutritional, Technological, Antioxidant, and Sensory Properties.

Grape pomace powder (GPP), a by-product from the winemaking process, was used to substitute flour for wheat bread fortification within 0, 5, and 10 g/100 g. Rheological properties of control and fortified doughs, along with physicochemical and nutritional characteristics, antioxidant activity, and the sensory analysis of the obtained bread were considered. The GPP addition influenced the doughs' rheological properties by generating more tenacious and less extensible products. Concerning bread, pH values and volume of fortified products decreased as the GPP inclusion level increased in the recipe. Total phenolic compounds and the antioxidant capacity of bread samples, evaluated by FRAP (ferric reducing ability of plasma) and ABTS (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)) assays, increased with GPP addition. Moreover, the GPP inclusion level raised the total dietary fiber content of bread. Regarding sensory evaluation, GPP fortification had a major impact on the acidity, the global flavor, the astringency, and the wine smell of bread samples without affecting the overall bread acceptability. The current results suggest that GPP could be an attractive ingredient used to obtain fortified bread, as it is a source of fiber and polyphenols with potentially positive effects on human health.

Monitoring the antioxidant activity of an eco-friendly processed grape pomace along the storage.

The aims of this research are to compare and to monitor two conditions for preserving the total phenolic content (TPC) and the antioxidant activity (AOA) of grape pomace (GP) processed as powder and its corresponding extract at room and freezing temperature, respectively. The highest TPC and AOA were obtained in the GP extracted in a ratio 1:10 (w/v) with ethanol at 50% for 45 min at 50 °C. After 9 months of room temperature (RT) storage, the GP powder obtained a significantly higher AOA than the initial condition and the extract frozen-stored the same time.

Impact of Grape Pomace Powder on the Phenolic Bioaccessibility and on In Vitro Starch Digestibility of Wheat Based Bread.

Breads were prepared by substituting common wheat flour with 0 (GP0), 5 (GP5) and 10 (GP10) g/100 g (w/w) of grape pomace powder (GPP) and were analyzed for the phenolic profile bioaccessibility as well as the in vitro starch digestion during simulated digestion. The free and bound phenolic composition of native GPP and resulting breads were profiled using ultra-high-performance chromatography-quadrupole-time-of-flight (UHPLC-QTOF). The raw GPP was characterized by 190 polyphenols with the anthocyanins representing the most abundant class, accounting for 11.60 mg/g of cyanidin equivalents. Regarding the fortified bread, the greatest (p < 0.05) content in phenolic compounds was recorded for the GP10 sample (considering both bound and free fractions) being 127.76 mg/100 g dry matter (DM), followed by the GP5 (106.96 mg/100 g DM), and GP0 (63.76 mg/100 g DM). The use of GPP determined an increase of anthocyanins (considered the markers of the GPP inclusion), recording 20.98 mg/100 g DM in GP5 and 35.82 mg/100 g DM in GP10. The bioaccessibility of anthocyanins increased in both GP5 and GP10 breads when moving from the gastric to the small intestine in vitro digestion phase with an average value of 24%. Both the starch hydrolysis and the predicted glycemic index decreased with the progressive inclusion of GPP in bread. Present findings showed that GPP in bread could promote an antioxidant environment in the digestive tract and influence the in vitro starch digestion.

Effect of Moringa oleifera L. Leaf Powder Addition on the Phenolic Bioaccessibility and on In Vitro Starch Digestibility of Durum Wheat Fresh Pasta.

Fresh pasta was formulated by replacing wheat semolina with 0, 5, 10, and 15 g/100 g (w/w) of Moringa oleifera L. leaf powder (MOLP). The samples (i.e., M0, M5, M10, and M15 as a function of the substitution level) were cooked by boiling. The changes in the phenolic bioaccessibility and the in vitro starch digestibility were considered. On the cooked-to-optimum samples, by means of ultra-high-performance liquid chromatography-quadrupole time-of-flight (UHPLC-QTOF) mass spectrometry, 152 polyphenols were putatively annotated with the greatest content recorded for M15 pasta, being 2.19 mg/g dry matter (p < 0.05). Multivariate statistics showed that stigmastanol ferulate (VIP score = 1.22) followed by isomeric forms of kaempferol (VIP scores = 1.19) and other phenolic acids (i.e., schottenol/sitosterol ferulate and 24-methylcholestanol ferulate) were the most affected compounds through the in vitro static digestion process. The inclusion of different levels of MOLP in the recipe increased the slowly digestible starch fractions and decreased the rapidly digestible starch fractions and the starch hydrolysis index of the cooked-to-optimum samples. The present results showed that MOLP could be considered a promising ingredient in fresh pasta formulation.

Effects of wheat germ agglutinin on human gastrointestinal epithelium: insights from an experimental model of immune/epithelial cell interaction.

Wheat germ agglutinin (WGA) is a plant protein that binds specifically to sugars expressed, among many others, by human gastrointestinal epithelial and immune cells. WGA is a toxic compound and an anti-nutritional factor, but recent works have shown that it may have potential as an anti-tumor drug and as a carrier for oral drugs. To quantitate the toxicity threshold for WGA on normal epithelial cells we previously investigated the effects of the lectin on differentiated Caco2 cells, and showed that in the micromolar range of concentrations WGA could alter the integrity of the epithelium layer and increase its permeability to both mannitol and dextran. WGA was shown to be uptaken by Caco2 cells and only approximately 0.1% molecules were observed to cross the epithelium layer by transcytosis. Here we show that at nanomolar concentrations WGA is unexpectedly bioactive on immune cells. The supernatants of WGA-stimulated peripheral blood mononuclear cells (PBMC) can alter the integrity of the epithelium layer when administered to the basolateral side of differentiated Caco2 cells and the effects can be partially inhibited by monoclonal antibodies against IL1, IL6 and IL8. At nanomolar concentrations WGA stimulates the synthesis of pro-inflammatory cytokines and thus the biological activity of WGA should be reconsidered by taking into account the effects of WGA on the immune system at the gastrointestinal interface. These results shed new light onto the molecular mechanisms underlying the onset of gastrointestinal disorders observed in vivo upon dietary intake of wheat-based foods.

Effects of microencapsulation by ionic gelation on the oxidative stability of flaxseed oil.

Flaxseed oil is a major source of omega-3 polyunsaturated fatty acids (PUFAs), as it contains nearly 50% of alpha-linolenic acid. For this reason it is highly susceptible to auto-oxidation. The aim of the work was to increase the stability of flaxseed oil by a microencapsulation process based on ionic gelation through vibrating-nozzle extrusion technology, using pectin as shell material. Two different drying systems, passive air drying (AD) and fluid bed (FB), were compared. The results show that the encapsulation efficiency is very high (up to 98%). Besides being approximately 20-fold faster, FB gives beads showing on average higher payload (76% vs 68%) and lower peroxide value (9.64 vs 21.33) than the AD. An accelerated test carried out on FB-dried beads shows that the oxidative stability of encapsulated oil is 13-fold higher than bulk oil (PV FB: 20 vs PV oil: 260), demonstrating the protecting effect of microencapsulation.

Isolation and identification of two lipid transfer proteins in pomegranate (Punica granatum).

Lipid transfer proteins (LTPs) are a family of low molecular mass (7-9 kDa) polypeptides, the members of which share 35-95% sequence homology. These proteins are widely distributed throughout the plant kingdom and are receiving attention for their biochemical characteristics and biological activity. LTPs are indeed studied in different research fields varying from allergy to food technology, and numerous molecules belonging to this class are progressively being identified and investigated. Proteins from pomegranate juice were fractioned by cation exchange chromatography and analyzed by SDS-PAGE. Two proteins were identified as putative LTPs on the basis of their molecular weights and their electrophoretic behaviors under reducing and nonreducing conditions. Finally, proteins were purified and characterized by mass spectrometry. This analysis confirmed that the two polypeptides are LTPs on the basis of an amino acid sequence common to LTPs from other plant sources and cysteine content. The two proteins, named LTP1a and LTP1b, showed similar molecular masses but different immunological profiles when immunodetected with rabbit antibodies specific for Pru p 3 and human IgE from a patient suffering from pomegranate allergy. The demonstration of the existence of two immunologically unrelated LTPs in pomegranate confirms the variability and the complexity of the plant LTP family. This should be taken into account when the role of these proteins as elicitors of allergies to fruits is investigated and could help to explain the contradictory literature data on pomegranate allergy.

Hen egg white lysozyme is a hidden allergen in Italian commercial ciders.

Hen egg white lysozyme (HEWL) is an enzyme used in alcoholic fermentation for its ability to control the growth of Gram-positive and spoilage bacteria, without inhibiting yeast growth, and it allows a reduction in the use of sulphur dioxide. Nevertheless, considering the potential allergenicity of this protein, the presence of HEWL should be declared on the label of the final product. In this work, we analysed 18 commercial Italian ciders by LC-MS/MS and found traces of HEWL in 12 samples without label declaration. We used Western blot and enzyme-linked immunosorbent assay (ELISA) to verify the immunological activity of HEWL, and to quantify its content in the ciders. Two out of 18 samples were found to be positive both by immunoblot and ELISA. The results indicate the requirement of a more stringent control of commercial ciders and the need of label declaration for ciders treated with such compounds.

Production of stable food-grade microencapsulated astaxanthin by vibrating nozzle technology.

Astaxanthin is a carotenoid known for its strong antioxidant and health-promoting characteristics, but it is also highly degradable and thus unsuited for several applications. We developed a sustainable method for the extraction and the production of stable astaxanthin microencapsulates. Nearly 2% astaxanthin was extracted by high-pressure homogenization of dried Haematococcus pluvialis cells in soybean oil. Astaxanthin-enriched oil was encapsulated in alginate and low-methoxyl pectin by Ca2+-mediated vibrating-nozzle extrusion technology. The 3% pectin microbeads resulted the best compromise between sphericity and oil retention upon drying. We monitored the stability of these astaxanthin beads under four different conditions of light, temperature and oxygen exposition. After 52weeks, the microbeads showed a total-astaxanthin retention of 94.1±4.1% (+4°C/-light/+O2), 83.1±3.2% (RT/-light/-O2), 38.3±2.2% (RT/-light/+O2), and 57.0±0.4% (RT/+light/+O2), with different degradation kinetics. Refrigeration, therefore, resulted the optimal storage condition to preserve astaxanthin stability.

Anti-tumour potential of a gallic acid-containing phenolic fraction from Oenothera biennis.

A phenolic fraction purified form defatted seeds of Oenothera biennis promoted selective apoptosis of human and mouse bone marrow-derived cell lines following first-order kinetics through a caspase-dependent pathway. In non-leukemia tumour cell lines, such as human colon carcinoma CaCo(2) cells and mouse fibrosarcoma WEHI164 cells, this fraction inhibited (3)H-thymidine incorporation but not cell death or cell cycle arrest. Human peripheral blood mononuclear cells showed low sensitivity to treatment. Single bolus injection of the phenolic fraction could delay the growth of established myeloma tumours in syngeneic animals. HPLC and mass spectrometry analysis revealed that the fraction contains gallic acid. However, the biological activity of the fraction differs from the activity of this phenol and hence it should be attributed to other co-purified molecules which remain still unidentified.

Studies on the joint cytotoxicity of Wheat Germ Agglutinin and monensin.

Wheat Germ Agglutinin (WGA) cytotoxicity has been studied using two human leukemia cell lines, Molt3 and K562, and human peripheral blood mononuclear cells (PBMC). In spite of similar binding at the cell surface, WGA was found to promote cell death to a different extent in Molt3, K562 and PBMC and to induce different death events leading to apoptosis in Molt3 and either apoptosis and necrosis in K562 cells and PBMC. In Molt3 but not in K562 cells, WGA cytotoxicity could be potentiated 66-200 fold by 50 nM monensin, a carboxylic ionophore that perturbs the intracellular trafficking of endocytosed molecules. Synergism between the cytotoxic activities of WGA and monensin was demonstrated in Molt3 cells by comparing non toxic, or slightly toxic, doses of WGA and monensin alone or in combination. These results show that the cytotoxic effect of WGA is dependent on internalisation events which may differ among the cell lines used. WGA and monensin can enter the human diet being a component of wheat germ and an antibiotic used for zootechnic reasons in the bioindustry, respectively. These data reveal the synergistic effect between two dietary molecules, otherwise per se toxic at much higher concentrations, with possible implications for human and animal health.

Quantitative determination of dietary lectin activities by enzyme-linked immunosorbent assay using specific glycoproteins immobilized on microtiter plates.

An immunoenzymatic method for the quantitative determination of dietary lectin activities employing immobilized glycoproteins was studied. Lectins from wheat germ (WGA), peanut (PNA), and jack bean (ConA) were added to microtiter plates coated with ovalbumin or asialofetuin and quantified by enzyme-linked immunosorbent assay (ELISA) with lectin-specific antibodies. ELISA responses for lectin activity were dose-dependent in the concentration range 30-1000 ng/mL for WGA and 80-1000 ng/mL for both PNA and ConA. Inhibition assays carried out with different saccharides confirmed that the binding of lectins to immobilized glycoproteins was specific. The proposed method is specific and sensitive, allowing the quantitative determination of lectin activities on raw samples by simple dilution of the extracts. Examples of application to wheat germ and roasted peanut extracts are reported.