• Cold Atmospheric Plasma Stimulates Clathrin-Dependent Endocytosis to Repair Oxidised Membrane and Enhance Uptake of Nanomaterial in Glioblastoma Multiforme Cells

      He, Zhonglei; Liu, Kangze; Scally, Laurence; Manaloto, Eline; Gunes, Sebnem; Ng, Sing Wei; Maher, Marcus; Tiwari, Brijesh K; Byrne, Hugh J.; Bourke, Paula; et al. (Springer Science and Business Media LLC, 2020-04-24)
      Cold atmospheric plasma (CAP) enhances uptake and accumulation of nanoparticles and promotes synergistic cytotoxicity against cancer cells. However, the mechanisms are not well understood. In this study, we investigate the enhanced uptake of theranostic nanomaterials by CAP. Numerical modelling of the uptake of gold nanoparticle into U373MG Glioblastoma multiforme (GBM) cells predicts that CAP may introduce a new uptake route. We demonstrate that cell membrane repair pathways play the main role in this stimulated new uptake route, following non-toxic doses of dielectric barrier discharge CAP. CAP treatment induces cellular membrane damage, mainly via lipid peroxidation as a result of reactive oxygen species (ROS) generation. Membranes rich in peroxidised lipids are then trafficked into cells via membrane repairing endocytosis. We confirm that the enhanced uptake of nanomaterials is clathrin-dependent using chemical inhibitors and silencing of gene expression. Therefore, CAP-stimulated membrane repair increases endocytosis and accelerates the uptake of gold nanoparticles into U373MG cells after CAP treatment. We demonstrate the utility of CAP to model membrane oxidative damage in cells and characterise a previously unreported mechanism of membrane repair to trigger nanomaterial uptake. This knowledge will underpin the development of new delivery strategies for theranostic nanoparticles into cancer cells.
    • Effect of high pressure processing on the safety, shelf life and quality of raw milk

      Stratakos, Alexandros Ch.; Inguglia, Elena Sofia; Linton, Mark; Tollerton, Joan; Murphy, Liam; Corcionivoschi, Nicolae; Koidis, Anastasios; Tiwari, Brijesh K (Elsevier, 2019-01-14)
      High pressure processing (HPP) was investigated as an alternative to standard raw milk processing. Different pressure levels (400–600 MPa) and exposure times (1–5 min) were tested against artificially inoculated pathogenic E. coli, Salmonella and L. monocytogenes. HPP effectively inactivated bacterial concentration by 5 log CFU/ml. The most effective HPP conditions in terms of pathogen reduction were subsequently utilised to determine the effect of pressure on microbiological shelf life, particle size and colour of milk during refrigerated storage. Results were compared to pasteurised and raw milk. HPP (600 MPa for 3 min) also significantly reduced the total viable counts, Enterobacteriaceae, lactic acid bacteria and Pseudomonas spp. in milk thus prolonging the microbiological shelf life of milk by 1 week compared to pasteurised milk. Particle size distribution curves of raw, pasteurised and HPP milk, showed that raw and HPP milk had more similar casein and fat particle sizes compared to pasteurised milk. The results of this study show the possibility of using HPP to eliminate pathogens present in milk while maintaining key quality characteristics similar to those of raw milk.
    • Fructooligosaccharides integrity after atmospheric cold plasma and high-pressure processing of a functional orange juice

      Lima Almeida, Francisca Diva; Gomes, Wesley Faria; Cavalcante, Rosane; Tiwari, Brijesh K; Cullen, Patrick J.; Frias, Jesus; Bourke, Paula; Fernandes, Fabiano A.N.; Rodrigues, Sueli; National Council of Technological and Scientific Development (Elsevier, 2017-10-02)
      In this study, the effect of atmospheric pressure cold plasma and high-pressure processing on the prebiotic orange juice was evaluated. Orange juice containing 7 g/100 g of commercial fructooligosaccharides (FOS) was directly and indirectly exposed to a plasma discharge at 70 kV with processing times of 15, 30, 45 and 60 s. For high-pressure processing, the juice containing the same concentration of FOS was treated at 450 MPa for 5 min at 11.5 °C in an industrial equipment (Hyperbaric, model: 300). After the treatments, the fructooligosaccharides were qualified and quantified by thin layer chromatography. The organic acids and color analysis were also evaluated. The maximal overall fructooligosaccharides degradation was found after high-pressure processing. The total color difference was < 3.0 for high-pressure and plasma processing. citric and ascorbic acid (Vitamin C) showed increased content after plasma and high-pressure treatment. Thus, atmospheric pressure cold plasma and high-pressure processing can be used as non-thermal alternatives to process prebiotic orange juice.
    • Future Protein Supply and Demand: Strategies and Factors Influencing a Sustainable Equilibrium

      Henchion, Maeve; Hayes, Maria; Mullen, Anne Maria; Fenelon, Mark; Tiwari, Brijesh K; Department of Agriculture, Food and the Marine, Ireland; 11/F/043 (MDPI, 20/07/2017)
      A growing global population, combined with factors such as changing socio-demographics, will place increased pressure on the world’s resources to provide not only more but also different types of food. Increased demand for animal-based protein in particular is expected to have a negative environmental impact, generating greenhouse gas emissions, requiring more water and more land. Addressing this “perfect storm” will necessitate more sustainable production of existing sources of protein as well as alternative sources for direct human consumption. This paper outlines some potential demand scenarios and provides an overview of selected existing and novel protein sources in terms of their potential to sustainably deliver protein for the future, considering drivers and challenges relating to nutritional, environmental, and technological and market/consumer domains. It concludes that different factors influence the potential of existing and novel sources. Existing protein sources are primarily hindered by their negative environmental impacts with some concerns around health. However, they offer social and economic benefits, and have a high level of consumer acceptance. Furthermore, recent research emphasizes the role of livestock as part of the solution to greenhouse gas emissions, and indicates that animal-based protein has an important role as part of a sustainable diet and as a contributor to food security. Novel proteins require the development of new value chains, and attention to issues such as production costs, food safety, scalability and consumer acceptance. Furthermore, positive environmental impacts cannot be assumed with novel protein sources and care must be taken to ensure that comparisons between novel and existing protein sources are valid. Greater alignment of political forces, and the involvement of wider stakeholders in a governance role, as well as development/commercialization role, is required to address both sources of protein and ensure food security.
    • Future Protein Supply and Demand: Strategies and Factors Influencing a Sustainable Equilibrium

      Henchion, Maeve; Hayes, Maria; Mullen, Anne Maria; Fenelon, Mark; Tiwari, Brijesh K; Department of Agriculture, Food and the Marine, Ireland (MDPI, 20/07/2017)
      A growing global population, combined with factors such as changing socio-demographics, will place increased pressure on the world’s resources to provide not only more but also different types of food. Increased demand for animal-based protein in particular is expected to have a negative environmental impact, generating greenhouse gas emissions, requiring more water and more land. Addressing this “perfect storm” will necessitate more sustainable production of existing sources of protein as well as alternative sources for direct human consumption. This paper outlines some potential demand scenarios and provides an overview of selected existing and novel protein sources in terms of their potential to sustainably deliver protein for the future, considering drivers and challenges relating to nutritional, environmental, and technological and market/consumer domains. It concludes that different factors influence the potential of existing and novel sources. Existing protein sources are primarily hindered by their negative environmental impacts with some concerns around health. However, they offer social and economic benefits, and have a high level of consumer acceptance. Furthermore, recent research emphasizes the role of livestock as part of the solution to greenhouse gas emissions, and indicates that animal-based protein has an important role as part of a sustainable diet and as a contributor to food security. Novel proteins require the development of new value chains, and attention to issues such as production costs, food safety, scalability and consumer acceptance. Furthermore, positive environmental impacts cannot be assumed with novel protein sources and care must be taken to ensure that comparisons between novel and existing protein sources are valid. Greater alignment of political forces, and the involvement of wider stakeholders in a governance role, as well as development/commercialization role, is required to address both sources of protein and ensure food security
    • Impact of pulsed electric field pre-treatment on nutritional and polyphenolic contents and bioactivities of light and dark brewer's spent grains

      Kumari, Bibha; Tiwari, Brijesh K; Walsh, Des; Griffin, Tomás; Islam, Nahidul; Lyng, James G.; Brunton, Nigel; Rai, Dilip K.; Department of Agriculture, Food and the Marine; European Union; et al. (Elsevier, 2019-04-30)
      Pulsed electric field (PEF) pre-treatment, at 2.8 kV/cm with 3000 pulses of 20 μs pulse-width, was applied on brewer's spent grains (BSG) followed by aqueous extraction at 55 °C, 220 rpm for 16 h. PEF pre-treatment showed significantly increased yields (p < 0.05) of carbohydrate, protein, starch and reducing sugar in extracts from dark BSG compared to untreated samples. Light BSG extracts had significantly higher (p < 0.05) levels of free d-glucose and total free amino acids (18.5–33.3 and 21–25 mg/g dry weight extract (Dwe)), compared to dark extracts (5 and 1.2 mg/g Dwe respectively). Dark BSG extracts showed significantly higher (p < 0.05) total phenolics (3.97–4.88 mg GAE/g Dwe) compared to light BSG extracts (0.83–1.40 mg GAE/g Dwe). Furthermore, PEF treated light BSG showed higher antimicrobial activity with minimum inhibition concentration (MIC) of 50 and 25 mg/mL against Salmonella typhimurium and Listeria monocytogenes, respectively compared to the untreated extracts (>50 mg/mL) with lowest MIC value of 1.56 mg/mL against Staphylococcus aureus. All the BSG extracts induced the release of pro-inflammatory cytokines (IL-1β, IL-6, TNF-α) and chemokines (IL-8, MCP-1 and MIP-1α) confirming immunomodulatory activity.
    • Integrated phenotypic-genotypic approach to understand the influence of ultrasound on metabolic response of Lactobacillus sakei

      Ojha, K. Shikha; Burgess, Catherine; Duffy, Geraldine; Kerry, Joseph P.; Tiwari, Brijesh K; Teagasc Walsh Fellowship Programme (PLOS, 2018-01-25)
      The lethal effects of soundwaves on a range of microorganisms have been known for almost a century whereas, the use of ultrasound to promote or control their activity is much more recent. Moreover, the fundamental molecular mechanism influencing the behaviour of microorganisms subjected to ultrasonic waves is not well established. In this study, we investigated the influence of ultrasonic frequencies of 20, 45, 130 and 950 kHz on growth kinetics of Lactobacillus sakei. A significant increase in the growth rate of L. sakei was observed following ultrasound treatment at 20 kHz despite the treatment yielding a significant reduction of ca. 3 log cfu/mL in cells count. Scanning electron microscopy showed that ultrasound caused significant changes on the cell surface of L. sakei culture with the formation of pores “sonoporation”. Phenotypic microarrays showed that all ultrasound treated L. sakei after exposure to various carbon, nitrogen, phosphorus and sulphur sources had significant variations in nutrient utilisation. Integration of this phenotypic data with the genome of L. sakei revealed that various metabolic pathways were being influenced by the ultrasound treatments. Results presented in this study showed that the physiological response of L. sakei in response to US is frequency dependent and that it can influence metabolic pathways. Hence, ultrasound treatments can be employed to modulate microbial activity for specialised applications.
    • Marine Functional Foods Research Initiative (NutraMara)

      Troy, Declan J.; Tiwari, Brijesh K; Hayes, Maria; Ross, R Paul; STANTON, CATHERINE; Johnson, Mark; Stengel, Dagmar; O'Doherty, John V.; Fitzgerald, Richard J.; McSorley, Emeir; et al. (Marine Institute, 2017-12)
      The NutraMara – Marine Functional Foods Research Initiative was conceived by Sea Change - A Marine Knowledge, Research and Innovation Strategy for Ireland 2007-2013. The goal was to develop a collaborative funding mechanism that would create new research capacity and build the capabilities required to maximise the potential of Ireland’s extensive marine bioresources. By supporting a strong interdisciplinary research team, capable of exploring marine animals and plants as a sustainable source of materials for use as functional ingredients and foods, the vision for NutraMara was to position Ireland to the fore in use of marine bioresources as health beneficial ingredients. Commencing in 2008 and supported by funds of €5.2 million from the Marine Institute and the Department of Agriculture, Food and the Marine, the research programme was led by Teagasc as the head of a multi-institutional consortium. The NutraMara consortium comprises marine bioresources and bioscience expertise, with food science and technology expertise from University College Cork; University College Dublin; the National University of Ireland Galway; the University of Limerick and Ulster University. Research effort was directed towards exploring Ireland’s marine bioresources – including macro- and microalgae, finfish and shellfish from wild and cultured sources: and discards from processing fish as sources of novel ingredients with bioactive characteristics. This discovery activity involved the collection of over 600 samples from 39 species of algae and fish and the analysis of 5,800 extracts, which resulted in 3,000 positive “hits” for bioactivity. The NutraMara consortium has built a strong research capacity to identify, characterise and evaluate marine-origin bioactives for use as/in functional foods. It further built the capacity to develop model foods enhanced with these marine-origin functional ingredients; providing insights to the processing challenges associated with producing functional ingredients from marine organisms. The consortium was actively engaged in research activities designed to identify and assess bioactive compounds from available marine resources, including polyphenols, proteins/peptides, amino acids, polysaccharides, polyunsaturated fatty acids and materials with antioxidant, probiotic or prebiotic properties. A key component of NutraMara’s activities was the development of human capital. The recruitment of M.Sc. and PhD students and their integration within a dynamic research environment that has strong links to industry, provided lasting expertise and capabilities, which are relevant to the needs of Ireland’s food and marine sectors. NutraMara research led to the awarding of eighteen PhDs and recruitment of 21 post-doctoral researchers over the eight year research programme. In excess of 80 peer reviewed publications resulted from this research and more publications are planned. A further 100 posters and conference presentations were also delivered by NutraMara researchers and Principal Investigators. The development and implementation of training and exchange programmes aimed at providing early stage researchers with inter-disciplinary skills that are critical to their development as researchers, enhanced the research capacity of institutions, the industry sectors and the country as a whole. Principal Investigators involved in leading the NutraMara research programme have secured additional research grants of almost €6 million from national and international sources and are engaged in extensive research collaboration involving marine and food research expertise; an activity which did not exist prior to NutraMara. The dissemination of knowledge and transfer of research results to industry were key activities in the research programme. The research outputs and visibility of NutraMara activity nationally resulted in 10 companies engaging in research and development activity with the consortium. Regular workshops and conferences organised by NutraMara attracted close to five hundred participants from Ireland and overseas. Members of the NutraMara core PI group have contributed to the formulation of new national foods and marine research policy and national research agenda, both during the national prioritisation exercise and in sectoral research strategies. This final project report describes the process by which research targets were identified, and the results of extensive screening and evaluation of compounds extracted from marine bioresources. It also highlights the development of new protocols designed to extract compounds in ways that are food friendly. Evaluating the functional properties, bioactivity and bioavailability of high potential marine compounds involved in vitro and in vivo testing. Pilot animal and human intervention studies yielded further insight to the potential and challenges in developing marine functional ingredients. As a result of work completed within the NutraMara consortium, Ireland is well positioned to continue to contribute to the development of ingredients derived from marine organisms and in doing so support the on-going development of Ireland’s food sector.
    • Optimisation of Ultrasound Frequency, Extraction Time and Solvent for the Recovery of Polyphenols, Phlorotannins and Associated Antioxidant Activity from Brown Seaweeds

      Ummat, Viruja; Tiwari, Brijesh K; Jaiswal, Amit K; Condon, Kevin; Garcia-Vaquero, Marco; O’Doherty, John; O’Donnell, Colm; Rajauria, Gaurav; Science Foundation Ireland; 16/RC/3889; et al. (MDPI AG, 2020-05-11)
      This study investigates ultrasound assisted extraction (UAE) process parameters (time, frequency and solvent) to obtain high yields of phlorotannins, flavonoids, total phenolics and associated antioxidant activities from 11 brown seaweed species. Optimised UAE conditions (35 kHz, 30 min and 50% ethanol) significantly improved the extraction yield from 1.5-fold to 2.2-fold in all seaweeds investigated compared to solvent extraction. Using ultrasound, the highest recovery of total phenolics (TPC: 572.3 ± 3.2 mg gallic acid equivalent/g), total phlorotannins (TPhC: 476.3 ± 2.2 mg phloroglucinol equivalent/g) and total flavonoids (TFC: 281.0 ± 1.7 mg quercetin equivalent/g) was obtained from Fucus vesiculosus seaweed. While the lowest recovery of TPC (72.6 ± 2.9 mg GAE/g), TPhC (50.3 ± 2.0 mg PGE/g) and TFC (15.2 ± 3.3 mg QE/g) was obtained from Laminaria digitata seaweed. However, extracts from Fucus serratus obtained by UAE exhibited the strongest 1,1-diphenyl-2-picryl-hydrazyl (DPPH) scavenging activity (29.1 ± 0.25 mg trolox equivalent/g) and ferric reducing antioxidant power (FRAP) value (63.9 ± 0.74 mg trolox equivalent/g). UAE under optimised conditions was an effective, low-cost and eco-friendly technique to recover biologically active polyphenols from 11 brown seaweed species.
    • Optimised protein recovery from mackerel whole fish by using sequential acid/alkaline isoelectric solubilization precipitation (ISP) extraction assisted by ultrasound

      Álvarez García, Carlos; Lélu, Pauline; Lynch, Sarah A.; Tiwari, Brijesh K; National Development Plan 2007–2013; MFFRI/07/01 (Elsevier, 2017-10-04)
      The growing fishery industry needs to find new green-processes in order to provide a solution to the huge amount of wastes and by-products that such industrial activity produces. Currently, around a 40% of the total weight of the mackerel is considered a by-product, because just the fillets are used in the food market. ISP method has been revealed as a useful tool for protein recovering, however the yield of this process is traditionally lower than enzymatic methods. In present work, the use of sequential acid/alkaline extraction and alkaline extraction assisted by ultrasound, have been implemented in order to increase the yield of the process. It has been demonstrated that (i) sequential extraction is able to recover practically 100% of total protein, and (ii) applying ultrasound to alkaline extraction is possible to recover more than 95% of total protein from mackerel by-products. Extracted proteins were characterized according to their size, and the amino acid profile of final product was determined.
    • Survival characteristics of monophasic Salmonella Typhimurium 4,[5],12:i:- strains derived from pig feed ingredients and compound feed

      Burns, Ann Marie; Duffy, Geraldine; Walsh, Des; Tiwari, Brijesh K; Grant, Jim; Lawlor, Peadar G; Gardiner, Gillian E.; Teagasc Walsh Fellowship Programme; 2011010 (Elsevier, 2015-12-09)
      The presence of Salmonella in animal feed or feed ingredients at the feed mill or on-farm is a cause for concern, as it can be transmitted to food-producing animals and subsequently to humans. The objective of this study was to determine the survival characteristics of five feed ingredient- and feed-derived monophasic Salmonella Typhimurium 4,[5],12:i:- strains. The first part of the study investigated thermal inactivation using an immersed heating coil apparatus. A Weibull model provided a good fit, with low RMSE values (0.04–0.43) and high R2 values (0.93–0.99) obtained. There was considerable inter-strain variation in heat resistance, with D-values ranging from 397.83 to 689 s at 55 °C, 11.35–260.95 s at 60 °C and 1.12 to 6.81 at 65 °C. Likewise, z-values ranged from 2.95 to 5.44 °C. One strain demonstrated a significantly higher thermal tolerance, even though it had been isolated from a meal feed. However, overall the strains investigated do not appear to be that much more heat resistant than Salmonella previously studied. The second part of this study involved assessing the ability of the five Salmonella strains to survive during storage over a 28-day period in pelleted weaner pig feed treated with 0.3% sodium butyrate. While a mean reduction in the Salmonella count of 0.79 log10 CFU was seen in the treated feed during the storage period, a reduction (albeit only 0.49 log10 CFU) was also observed in the control feed. Although there was no overall effect of treatment, sodium butyrate resulted in reductions in Salmonella counts of 0.75 and 0.22 log10 CFU at days 14 and 24 of feed storage, respectively but at the end of the 28-day storage period counts were 0.25 log10 CFU higher in the treated feed. Therefore, the sodium butyrate used appears unsuitable as an agent for feed treatment perhaps due to the protective coating on the particular feed additive used. Overall, the results of this study enhance knowledge about the behaviour and survival characteristics of monophasic S. Typhimurium 4,[5],12:i:- strains in animal feed and may assist the feed industry and pig producers in implementing effective intervention strategies for their control.
    • Ultrasound-assisted extraction of polyphenols from potato peels: profiling and kinetic modelling

      Kumari, Bibha; Tiwari, Brijesh K; Hossain, Mohammad Billal; Rai, Dilip K.; Brunton, Nigel; Department of Agriculture, Food and Marine; FIRM/11/F/050 (Wiley, 2017-02-14)
      Ultrasound‐assisted extraction (UAE) at 33 and 42 kHz has been investigated in the extraction of polyphenols from peels of two potato varieties, cream‐skinned Lady Claire (LC) and pink‐skinned Lady Rosetta (LR), commonly used in snack food production. Extraction efficacy between the UAE‐untreated (control) and the UAE‐treated extracts was assessed on the total phenolic content and antioxidant capacities (DPPH and FRAP). Application of UAE showed significantly higher recovery of phenolic compounds compared to solid–liquid extraction process alone. Lower ultrasonic frequency (33 kHz) was more effective in recovering polyphenols compared to 42 kHz ultrasonic treatment. The liquid chromatography‐tandem mass spectrometry revealed that chlorogenic acid and caffeic acid were the most prevalent phenolics in LR peels, whereas caffeic acid was dominant in LC peels. Peleg's equation showed a good correlation (R2 > 0.92) between the experimental values and the predicted values on the kinetics of UAE of phenolic compounds.