HPLC-MS/MS Phenolic Characterization of Olive Pomace Extracts Obtained Using an Innovative Mechanical Approach.

Olive pomace results from the production of olive oil. Even if olive pomace represents a potential environmental problem, it contains phenolic compounds, which are widely recognized for their beneficial properties for human health. In this study, an innovative and sustainable technological approach to extract phenolic compounds from fresh olive pomace, based on food-grade solvent instead of those usually adopted, is investigated. Characterization and shelf-life evaluation of the hydroalcoholic extracts obtained from the procedure developed for different industrial purposes were also carried out. The phenolic fractions of the different samples were studied with the Folin-Ciocâlteu method to quantify that the total reducing molecules and HPLC-MS/MS analysis was used to define the profile through the identification and quantification of 42 compounds, belonging to five chemical families. Regarding shelf-life, the hydroalcoholic extract showed no significant reduction in phenolic content, for both instrumental evaluations, retaining most of the phenolic compounds present in the raw material; negative attributes were not perceived by sensory evaluation. Thus, these lab-scale results can be the starting point to develop a procedure that is suitable for a real olive mill, representing a valorization strategy in a circular economy and the perspective of new business models.

Future Perspective and Technological Innovation in Cheese Making Using Artichoke (Cynara scolymus) as Vegetable Rennet: A Review.

Milk coagulation is a process used for the formulation of different dairy products such as cheese. In this process, milk undergoes changes in its chemical stability thanks to acidification or enzymatic reactions. Traditionally, milk coagulation has been carried out with rennet of animal origin, but recently, the research of new types of rennet such as microbial rennet and vegetable rennet has increased. This study aims to present an organized review of the most relevant information on lactic coagulation, its relationship with vegetable rennets, and the importance of the botanical genus Cynara in the extraction of vegetable rennets, focusing on the coagulant potential of artichoke (Cynara scolymus). We conducted this literature review and found that lactic coagulation and vegetable rennets are linked through the enzymatic activity of the latter. The results of the main studies demonstrated a strong relationship between vegetable rennets and protease enzymes as well as the presence of these enzymes in extracts of cardoon (Cynara scolymus) and artichoke (Cynara scolymus). In addition, studies highlight the presence of thistle extracts in artisanal cheese preparations in the Iberian Peninsula. Based on the results of the studies, a comparison between cheeses made with vegetable rennet and those made with traditional rennet was also carried out. Although the results show that the use of vegetable rennet in the manufacture of cheese can confer undesirable characteristics, the use of extracts from Cynara plants demonstrates that vegetable rennets have an industrial potential, especially the one obtained from artichoke (Cynara scolymus) due to its high availability. Nevertheless, specific studies are required for a better understanding and application of this rennet.

Lipid Process Markers of Durum Wheat Debranning Fractions.

At present, whole grains are usually obtained by adding bran and middlings to refined flours, and this recombination leads to certain variations in the ratio of endosperm, bran and germ, resulting in flours with very different compositional characteristics and rheological properties. Therefore, this study focuses on the identification of specific lipid markers in different debranning fractions of Italian and Canadian durum wheat blends. The by-products obtained from five different debranning levels (3, 6, 9, 12 and 15%) had a high content of monounsaturated fatty acids and a higher concentration of tocopherols and sterols than the corresponding debranned grains. The Italian and Canadian durum wheat samples did not show significant differences in the content of these bioactive lipid compounds. In particular, palmitic acid, oleic acid, tocopherol isomers and total sterols could be useful biomarkers for evaluating the grain-to-tissue ratio in recombined flours.

When Cannabis sativa L. Turns Purple: Biosynthesis and Accumulation of Anthocyanins.

Environmental cues elicit anthocyanin synthesis in plant vegetative and reproductive tissues. Their accumulation in different organs accounts for their diverse biological functions, mainly related to their antioxidant properties, and it depends on a temporally and spatially regulated mechanism controlled by the action of a well-known multi-transcription factor complex. Despite the highly recognizable value of Cannabis sativa L. as a natural biorefinery of phytochemicals, very little information is known on anthocyanin pigmentation in this species. In this work, a targeted quantification of anthocyanins via HPLC-MS/MS, combined with the transcriptional profile via RT-qPCR of genes encoding for structural and decorating enzymes and regulatory transcription factors in different C. sativa tissues, help gain insights into the anthocyanin pathway in this species. To the best of our knowledge, this is the first report on the identification of cyanidin-3-rutinoside (keracyanin) as the major anthocyanin in C. sativa vegetative and floral tissues. Keracyanin amounts were higher than in small berries, suggesting that Cannabis biomass is a valuable source of colored antioxidants to be exploited in diverse applications. Furthermore, a gene putatively encoding for an anthocyanin DTX35 type transporter and CsTTG1 were identified in silico and their transcriptional levels were assessed via RT-qPCR. The results allow us to provide the first model of anthocyanin regulation in C. sativa, opening a new research scenario in this species for both breeding purposes and phytochemical exploitation.

Effect of Sprouting on Biomolecular and Antioxidant Features of Common Buckwheat (Fagopyrum esculentum).

Buckwheat is a pseudo-cereal widely grown and consumed throughout the world. Buckwheat is recognized as a good source of nutrients and, in combination with other health-promoting components, is receiving increasing attention as a potential functional food. Despite the high nutritional value of buckwheat, a variety of anti-nutritional features makes it difficult to exploit its full potential. In this framework, sprouting (or germination) may represent a process capable of improving the macromolecular profile, including reducing anti-nutritional factors and/or synthesizing or releasing bioactives. This study addressed changes in the biomolecular profile and composition of buckwheat that was sprouted for 48 and 72 h. Sprouting increased the content of peptides and free-phenolic compounds and the antioxidant activity, caused a marked decline in the concentration of several anti-nutritional components, and affected the metabolomic profile with an overall improvement in the nutritional characteristics. These results further confirm sprouting as a process suitable for improving the compositional traits of cereals and pseudo-cereals, and are further steps towards the exploitation of sprouted buckwheat as a high-quality ingredient in innovative products of industrial interest.

Assessment of Lipid Quality in Commercial Omega-3 Supplements Sold in the French Market.

Supplementation of omega-3 fatty acids is considered a valuable strategy to supply the low intake of these fatty acids. Thus, the safety of the supplements is an important milestone. Because of that, we analyzed 20 unflavored supplements sold in the French market for fatty acid and triglyceride composition, for EPA and DHA, and for tocol content, as well as for oxidative status. This study found that only 2.5% of the supplements did not meet their label claims for omega-3 content. TAG analysis showed high variability among the triglyceride distribution, and the same trend was also noticed for the tocol content; in fact, a high variability of the distribution of the six tocols (four tocopherols and two tocotrienols) was found among the samples. Of the tested products, all of them complied with peroxide value, p-anisidine value, and Totox values established by the Global Organization for EPA and DHA Omega-3s (GOED) and were not oxidized.

Co-milling process of olives and oleaginous matrices with high nutritional value: a preliminary characterisation of the obtained oils.

Extra virgin olive oil (EVOO), grape seeds (GS) and pomegranate seeds (PS) are very popular for human consumption because of their nutraceutical properties. A co-milling of olives with GS or PS was carried out with the aim of a preliminary study of the lipidic and phenolic characteristics of the obtained vegetable oils, also during their shelf life. Results show that the use of GS and PS in the olive co-milling enriches the oil in healthy compounds, such as punicic acid and γ-tocopherol. However, the co-milling process must be re-evaluated, since the compositional profile of the co-milled oils is just slightly different from EVOO. The oxidative state of the oils obtained suggests the use of a protective packaging combined with small containers in order to ensure a rapid consumption. With an appropriate formulation the co-milled oils could bring health-positive compounds and, simultaneously, raw materials could be valorised.

Antioxidant and Antimicrobial Activity of Hydroethanolic Leaf Extracts from Six Mediterranean Olive Cultivars.

Phenolic profiles, antioxidant, and antimicrobial activities of hydroethanolic olive leaf extracts from six Mediterranean olive cultivars (Croatian: Lastovka, Levantinka, Oblica; Italian: Moraiolo, Frantoio, Nostrana di Brisighella) were investigated. As expected, various distributions of phenolic levels were observed for each cultivar and the total phenolic content showed high variability (ranging from 4 to 22 mg GAE/g of dry extract), with the highest amount of phenolics found in the Oblica sample, which also provided the highest antiradical (ORAC) and reducing activity (FRAP). The screening of individual compounds was performed by HPLC-PDA-ESI-QTOF-MS and the main detected compounds were oleuropein, hydroxytyrosol, oleoside/secologanoside, verbascoside, rutin, luteolin glucoside, hydroxyoleuropein, and ligstroside. While the antioxidant activity of the samples was relatively high, they showed no bactericidal and bacteriostatic activity against E. coli and S. Typhimurium; weak activity against Staphylococcus aureus, Bacillus cereus, and Listeria innocua; and inhibitory effects against Campylobacter jejuni at 0.5 mg dry extract/mL. The obtained results support the fact that olive leaf extracts, and especially those from the Oblica cultivar, could potentially be applied in various industries as natural preservatives and effective and inexpensive sources of valuable antioxidants.

Wheat Germ and Lipid Oxidation: An Open Issue.

Wheat germ (WG)'s shelf life after the milling process is incredibly short because of the presence of enzymes that aggravate the oxidation process; thus, stabilization is required in order to exploit the nutrients and bioactive compounds within WG. The critical point for the oxidation process is the mechanical treatment used to separate WG from the kernel, which exposes the lipid fraction to the air. Showing the connection between the quality of durum wheat, considering its storage management, and wheat germ oil (WGO), extracted with a cold press, solvent and supercritical CO2, is the aim of the study. The acidity and peroxide values were analyzed to evaluate lipid oxidation, while fatty acids, tocols, sterols and policosanols were evaluated for WGO characterization. The first fundamental step to control lipid oxidation is raw material management. Subsequently, the tempering phase of durum wheat, which is applied before the degermination process, is the most critical point for oxidation to develop because of the increase in moisture in the caryopsis and the activation of lipase and lipoxygenase. This represents a paradox: in order to stabilize the germ with degermination, first it seems inevitable to carry out a process that destabilizes it. To retains its highest quality, this will lead to a better use of the whole grain by reducing WG and by-product waste.

Influence of different baking powders on physico-chemical, sensory and volatile compounds in biscuits and their impact on textural modifications during soaking.

The aim of this study was to investigate the effects of different type of leavening agents (ammonium bicarbonate-A, cream of tartar-B, grape-based leavening agent-C, baking powder-D) on the structural (also after soaking), physico-chemical and other quality characteristics of biscuits made with the same formulation. The major changes observed between samples were related to their textural properties, volatile profile and sensory characteristics. The presence of ammonium bicarbonate in sample A, gave arise to a biscuit with the most homogeneous surface appearance, the less hard structure and highest fracturability and spread, that promoted their greater milk uptake during soaking. The B and C biscuits, made with baking powder having tartaric acid, were characterized by a crisper texture and lower sensory overall acceptability. The presence in formulation of baking powders made with cream of tartar (B) and ammonium bicarbonate (A) mostly influenced the biscuit volatile profiles. Biscuit sample obtained with the most common baking powder (diphosphates powder) showed in general intermediate characteristics between A and B-C samples. Results obtained in this study demonstrate that a lot of final properties of biscuits can be strongly influenced by the type of leavening agents used in formulation.

Olive oil by-product as functional ingredient in bakery products. Influence of processing and evaluation of biological effects.

Nowadays, the strong demand for adequate nutrition is accompanied by concern about environmental pollution and there is a considerable emphasis on the recovery and recycling of food by-products and wastes. In this study, we focused on the exploitation of olive pomace as functional ingredient in biscuits and bread. Standard and enriched bakery products were made using different flours and fermentation protocols. After characterization, they were in vitro digested and used for supplementation of intestinal cells (Caco-2), which underwent exogenous inflammation. The enrichment caused a significant increase in the phenolic content in all products, particularly in the sourdough fermented ones. Sourdough fermentation also increased tocol concentration. The increased concentration of bioactive molecules did not reflect the anti-inflammatory effect, which was modulated by the baking procedure. Conventionally fermented bread enriched with 4% pomace and sourdough fermented, not-enriched bread had the greatest anti-inflammatory effect, significantly reducing IL-8 secretion in Caco-2 cells. The cell metabolome was modified only after supplementation with sourdough fermented bread enriched with 4% pomace, probably due to the high concentration of tocopherol that acted synergistically with polyphenols. Our data highlight that changes in chemical composition cannot predict changes in functionality. It is conceivable that matrices (including enrichment) and processing differently modulated bioactive bioaccessibility, and consequently functionality.

Distribution of Free and Bound Phenolic Compounds in Buckwheat Milling Fractions.

Buckwheat is a rich source of phenolic compounds that have shown to possess beneficial effect to reduce some diseases due to their antioxidant power. Phenolic compounds are present in the free and in the bound form to the cell wall that are concentrated mainly in the outer layer (hull and bran). Hull is removed before the milling of buckwheat to obtain flours. In order to evaluate the phenolic composition in dehulled buckwheat milling fractions, it was carried out a determination of free and bound phenolic compounds in dehulled whole buckwheat flour, light flour, bran flour, and middling flour by high-performance liquid chromatography-mass spectrometry (HPLC-MS). The most abundant free phenolic compounds were rutin and epiafzelchin-epicatechin-O-dimethylgallate, whereas the most abundant bound phenolic compounds were catechin and epicatechin in all buckwheat flours. Besides, the highest content of free phenolic compounds was obtained in bran flour (1249.49 mg/kg d.w.), whereas the greatest bound phenolic content was in middling (704.47 mg/kg d.w.) and bran flours (689.81 mg/kg d.w.). Thus, middling and bran flours are naturally enriched flours in phenolic compounds that could be used to develop functional foods.

Use of Sieving As a Valuable Technology to Produce Enriched Buckwheat Flours: A Preliminary Study.

Fractionation processes based on physical separation are a good strategy to produce enriched cereal flours. Therefore, the aim of this work is to evaluate the suitability of sieving of buckwheat flours to produce protein and phenolic (especially rutin) enriched fractions. Because of that, dehulled whole buckwheat flour (GSTQ) was sieved obtaining fractions with a particle size of 215 µm, 160 µm, 85 µm, and 45 µm (GS215, GS160, GS85, and GS45). For that purpose, the determination of protein, ash, and total starch content and free and bound phenolic compounds was carried out. The highest content of total phenolic compounds was obtained in GS215 (3118.84 mg Kg-1 d.w.), followed by GS160 (2499.11 mg Kg-1 d.w.), GS85 (989.46 mg Kg-1 d.w.), GSTQ (983.15 mg Kg-1 d.w.), and GS45 (481.31 mg Kg-1 d.w.). Therefore, the phenolic content decreased with the particle size decrease from 215 µm to 45 µm. Besides, there were no significant differences between the total phenolic content in GS85 and GSTQ. The fraction with 215 µm reported the highest protein and mineral salt content and presented rutin amounts four times higher than GSTQ.

Optimization of Sonotrode Ultrasonic-Assisted Extraction of Proanthocyanidins from Brewers' Spent Grains.

Brewing spent grains (BSGs) are the main by-product from breweries and they are rich of proanthocyanidins, among other phenolic compounds. However, literature on these compounds in BSGs is scarce. Thus, this research focuses on the establishment of ultrasound-assisted extraction of proanthocyanidin compounds in brewing spent grains using a sonotrode. To set the sonotrode extraction up, response surface methodology (RSM) was used to study the effects of three factors, namely, solvent composition, time of extraction, and ultrasound power. Qualitative and quantitative analyses of proanthocyanidin compounds were performed using HPLC coupled to fluorometric and mass spectrometer detectors. The highest content of proanthocyanidins was obtained using 80/20 acetone/water (v/v), 55 min, and 400 W. The established method allows the extraction of 1.01 mg/g dry weight (d.w.) of pronthocyanidins from BSGs; this value is more than two times higher than conventional extraction.

(Ultra) High Pressure Homogenization Potential on the Shelf-Life and Functionality of Kiwifruit Juice.

The increasing competition within the food industry sector makes the requisite of innovation in processes and products essential, leading to focus the interest on the application of new processing technologies including high pressure homogenization (HPH) and ultra high pressure homogenization (UHPH). In this context, the present research aimed at evaluating the effects of two UHPH treatments performed at 200 MPa for 2 and 3 cycles on quality and functionality of organic kiwifruit juice stored at three different temperatures, i.e., 5, 15, and 25°C. The results showed that only the treatment performed at 200 MPa for 3 cycles was able to significantly increase the shelf-life of organic kiwifruit juices when stored at refrigeration temperature, avoiding also phase separation that occurred in the sample treated at 0.1 MPa (control) after 20 days of refrigerated storage. The obtained data showed also that the highest applied pressure was able to increase some quality parameters of the juice such as viscosity and luminosity (L∗) and increased the availability of total phenol content consequently enhancing the juice total antioxidant activity. The application of a treatment at 200 MPa for 3 cycles allowed to obtain a stable kiwifruit juice for more than 40 days under refrigerated storage. A challenge to implement this technology in food process as full alternative to thermal treatment could be represented by the adoption of pressure level up to 400 MPa followed by the packaging in aseptic conditions.

Recovery of Oligomeric Proanthocyanidins and Other Phenolic Compounds with Established Bioactivity from Grape Seed By-Products.

Grape seeds are a copious part of the grape pomace produced by wine and juice industry and they represent an interesting source of phenolic compounds. Proanthocyanidins (PAs) are the main class of grape seed phenols and are important dietary supplements for their well-known beneficial properties. In this study enriched extracts obtained from Chardonnay and Pignoletto grape seeds were characterized for their proanthocyanidins and other minor phenolic compounds content and composition. Seed PAs were fractionated using Sephadex LH-20, using different ethanol aqueous solutions as mobile phase and analysed by normal phase HPLC-FLD-ESI-MS. Monomers, oligomers up to dodecamers and polymers were recorded in all samples. For both cultivars, the extracts showed a high content in PAs. The determination of other phenolic compounds was carried out using a HPLC-QqQ-ESI-MS and Chardonnay samples reported a greater content compared to Pignoletto samples. Contrary to PAs fraction, extracts obtained with ethanol/water 50/50 (v/v) presented a significant higher phenolic content than the others.

Effect of Fermentation with Different Lactic Acid Bacteria Starter Cultures on Biogenic Amine Content and Ripening Patterns in Dry Fermented Sausages.

In the present study, two different diameter (small and large) Milano-type dry fermented sausages were industrially produced to evaluate the effect of two different LAB starter cultures (Lactobacillus sakei and Pediococcus pentosaceus) on biogenic amines (BAs) content, proteolysis, and lipolysis taking place during both fermentation and ripening. With regard to BAs, putrescine and tyramine were mostly found in fermented sausages having large diameter and those inoculated with P. pentosaceus/S. xylosus exhibited significantly higher accumulation of these compounds. Overall, the small size sausages showed a more pronounced proteolysis taking place during processing. In addition, aside from the distinctive electrophoretic bands detected with both starter cultures, a more pronounced proteolysis and a faster protein hydrolysis was observed in salami inoculated with P. pentosaceus/S. xylosus. As for lipolysis, a significantly higher amount of diacylglycerols was observed at the end of ripening in the sausages inoculated with L. sakei/S. xylosus, which concurrently exhibited an increased D32, D34, and D36 series. The results of the present study confirms profound differences in BAs concentration, proteolysis, and lipolysis. These findings are strictly dependent on the starter cultures, which demonstrates that the choice of an appropriate starter optimized for peculiar products and processes should be the key factor to improve safety and quality features of traditional fermented sausages.

Olive oil industry by-products. Effects of a polyphenol-rich extract on the metabolome and response to inflammation in cultured intestinal cell.

Over the past years, researchers and food manufacturers have become increasingly interested in olive polyphenols due to the recognition of their biological properties and probable role in the prevention of various diseases such as inflammatory bowel disease. Olive pomace, one of the main by-products of olive oil production, is a potential low-cost, phenol-rich ingredient for the formulation of functional food. In this study, the aqueous extract of olive pomace was characterized and used to supplement human intestinal cell in culture (Caco-2). The effect on the cell metabolome and the anti-inflammatory potential were then evaluated. Modification in the metabolome induced by supplementation clearly evidenced a metabolic shift toward a "glucose saving/accumulation" strategy that could have a role in maintaining anorexigenic hormone secretion and could explain the reported appetite-suppressing effect of the administration of polyphenol-rich food. In both basal and inflamed condition, supplementation significantly reduced the secretion of the main pro-inflammatory cytokine, IL-8. Thus, our data confirm the therapeutic potential of polyphenols, and specifically of olive pomace in intestinal bowel diseases. Although intervention studies are needed to confirm the clinical significance of our findings, the herein reported results pave the road for exploitation of olive pomace in the formulation of new, value-added foods. In addition, the application of a foodomics approach allowed observing a not hypothesized modulation of glucose metabolism.

Fermented Nut-Based Vegan Food: Characterization of a Home made Product and Scale-Up to an Industrial Pilot-Scale Production.

Because of the impossibility to consume food of animal origin, vegan consumers are looking for substitutes that could enrich their diet. Among many substitutes, fermented nut products are made from different nut types and obtained after soaking, grinding, and fermentation. Although other fermented vegetable products have been deeply investigated, there are few data about the fermentative processes of nut-based products and the microbial consortia able to colonize these products are not yet studied. This study characterized a hand-made vegan product obtained from cashew nut. Lactic acid bacteria responsible for fermentation were identified, revealing a succession of hetero- and homo-fermentative species during process. Successively, some lactic acid bacteria isolates from the home-made vegan product were used for a pilot-scale fermentation. The products obtained were characterized and showed features similar to the home-made one, although the microbiological hazards have been prevented through proper and rapid acidification, enhancing their safety features. PRACTICAL APPLICATION: Spontaneous fermented products are valuable sources of microorganisms that can be used in many food processes as starter cultures. The lactic acid bacteria isolated in this research can be exploited by industries to develop new foods and therefore to enter new markets. The use of selected starter cultures guarantees good organoleptic characteristics and food safety (no growth of pathogens).

Value-addition of Beef Meat By-products: Lipid Characterization by Chromatographic Techniques.

The lipid characterization of storage fat, subcutaneous fat and bone marrow, originated from three different bovine categories, calf, young bull and cow, was carried out in order to develop a re-use of these by-products. After the lipid extraction, the compositions in fatty acids, triacylglycerols and cholesterol were determined by GC-FID. A total of 25 fatty acids were identified in all by-products; the oleic acid was the preponderant component, followed by palmitic, stearic, palmitoleic, linoleic and myristic acid. The long chain triacylglycerols (T48, T50 and T52) were the main products, especially in the cow's by-products. The subcutaneous fat of all animals showed the highest cholesterol content, whereas it was present in low amount in the others. Phospholipids were analysed in bone marrow of all the animals and phosphatidylcholine was the most abundant compound. Because of the high fat content and essential fatty acids and the low cholesterol amount, storage fat and bone marrow could represent a valuable lipid resource in food and pharmaceutical industry.