Stabilisation of vitamin A by wheat bran is affected by wheat bran antioxidants, bound lipids and endogenous lipase activity.

Food fortification is an efficient strategy to combat vitamin A deficiency. However, the stability of vitamin A during storage is low. Cereal bran can be used as a natural and affordable stabilising agent, but the mechanism behind this stabilisation remains unclear. To unravel this mechanism, vitamin A stabilisation was studied during an accelerated storage experiment (60 °C, 70% relative humidity) using a set of 30 in-house modified wheat bran samples. The characteristics of these samples were linked to vitamin A stabilisation during storage using forward regression modelling. While all wheat bran samples could stabilise vitamin A to a significant extent, the stabilising effect was more pronounced for samples with a high antioxidant capacity, high bound lipid content and low lipase activity. The main effect of lipase activity was more than thrice as large as the main effects of antioxidant capacity and bound lipid content. These results suggest that wheat bran antioxidants and bound lipids protect vitamin A from degradation during storage, while endogenous lipase activity counteracts the stabilising effect. Based on these findings, modified wheat bran mixed with vitamin A can be a cost-effective and healthy aid in food fortification by providing high vitamin A stability.

Impact of process parameters on the specific volume of wholemeal wheat bread made using sourdough- and baker's yeast-based leavening strategies.

The final quality of wholemeal wheat bread is determined by the process parameter settings and leavening strategy. We hypothesise that the used leavening strategy may influence the optimal process parameter settings and, as such, the specific volume of the bread loaf. To analyse this interaction, bread was leavened with (i) a type 1 sourdough (SB), (ii) a type 1 sourdough combined with baker's yeast (YSB), or (iii) baker's yeast (YB). For each leavening strategy, the specific volume of bread, in response to variations in mixing time (4-10/4-14 min), water absorption (60-85 %), and proofing time (1-7/1-3 h), was analysed using an I-optimal response surface experimental design. Data modelling identified a substantially lower maximal specific volume of SB (2.13 mL/g), compared to YSB (3.30 mL/g) and YB (3.26 mL/g). The proofing time and water absorption mostly influenced the specific volume of the SB and YSB, respectively. However, the mixing and proofing times mainly affected the specific volume of YB. The type 1 sourdough reduced the mixing time and water absorption required for an optimal specific volume of bread compared to baker's yeast. These results challenge the idea of yielding higher volumes upon using sourdough compared to baker's yeast and highlight the importance of optimisation of bread dough formulations and breadmaking processes.

High-definition electroporation: Precise and efficient transfection on a microelectrode array.

Intracellular delivery is critical for a plethora of biomedical applications, including mRNA transfection and gene editing. High transfection efficiency and low cytotoxicity, however, are often beyond the capabilities of bulk techniques and synonymous with extensive empirical optimization. Moreover, bulk techniques are not amenable to large screening applications. Here, we propose an expeditious workflow for achieving optimal electroporation-based intracellular delivery. Using the multiplexing ability of a high-definition microelectrode array (MEA) chip, we performed a sequence of carefully designed experiments, multiple linear regression modelling and validation to obtain optimal conditions for on-chip electroporation of primary fibroblasts. Five electric pulse parameters were varied to generate 32 different electroporation conditions. The effect of the parameters on cytotoxicity and intracellular delivery could be evaluated with just two experiments. Most successful electroporation conditions resulted in no cell death, highlighting the low cytotoxicity of on-chip electroporation. The resulting delivery models were then used to achieve dosage-controlled delivery of small molecules, delivery of Cas9-GFP single-guide RNA complexes and transfection with an mCherry-encoding mRNA, resulting in previously unreported high-efficiency, single-cell transfection on MEAs: cells expressed mCherry on 81% of the actuated electrodes, underscoring the vast potential of CMOS MEA technology for the transfection of primary cells.

Superinfection exclusion factors drive a history-dependent switch from vertical to horizontal phage transmission.

Temperate bacterial viruses are commonly thought to favor vertical (lysogenic) transmission over horizontal (lytic) transmission when the virion-to-host-cell ratio is high and available host cells become scarce. In P22-infected Salmonella Typhimurium populations, however, we find that host subpopulations become lytically consumed despite high phage-to-host ratios that would normally favor lysogeny. These subpopulations originate from the proliferation of P22-free siblings that spawn off from P22-carrier cells from which they cytoplasmically inherit P22-borne superinfection exclusion factors (SEFs). In fact, we demonstrate that the gradual dilution of these SEFs in the growing subpopulation of P22-free siblings restricts the number of incoming phages, thereby imposing the perception of a low phage-to-host ratio that favors lytic development. Although their role has so far been neglected, our data indicate that phage-borne SEFs can spur complex infection dynamics and a history-dependent switch from vertical to horizontal transmission in the face of host-cell scarcity.

Robust dynamic experiments for the precise estimation of respiration and fermentation parameters of fruit and vegetables.

Dynamic models based on non-linear differential equations are increasingly being used in many biological applications. Highly informative dynamic experiments are valuable for the identification of these dynamic models. The storage of fresh fruit and vegetables is one such application where dynamic experimentation is gaining momentum. In this paper, we construct optimal O2 and CO2 gas input profiles to estimate the respiration and fermentation kinetics of pear fruit. The optimal input profiles, however, depend on the true values of the respiration and fermentation parameters. Locally optimal design of input profiles, which uses a single initial guess for the parameters, is the traditional method to deal with this issue. This method, however, is very sensitive to the initial values selected for the model parameters. Therefore, we present a robust experimental design approach that can handle uncertainty on the model parameters.

Feed endoxylanase type and dose affect arabinoxylan hydrolysis and fermentation in ageing broilers.

Despite the general use of endoxylanases in poultry feed to improve broiler performance, the abundance of different endoxylanase products and the variable response to their application in the field prevent a clear understanding of endoxylanase functionality in vivo. To gain insight into this functionality, we investigated the impact of endoxylanase type (Belfeed from Bacillus subtilis versus Econase XT from Nonomuraea flexuosa) and dose (10, 100, 1,000 mg/kg) in combination with broiler age on arabinoxylan (AX) hydrolysis and fermentation in broilers (Ross 308) fed a wheat-soy based diet. In a digestibility trial and a performance trial, a total of 1,057 one-day-old chicks received the control diet or 1 of the 6 endoxylanase supplemented wheat-soy based diets with, respectively, 5 replicate cages and 8 replicate pens per dietary treatment per trial. The AX content and structure, the AX digestibility values and the short-chain fatty acids produced were analysed at the level of the ileum, caeca and excreta at d 11 and 36. Endoxylanase supplementation resulted in a more extensive solubilisation of wheat AX and a reduction in the intestinal viscosity compared to the control (P < 0.05). A high endoxylanase dose was, however, required to obtain increased hydrolysis of the dietary AX along the gastrointestinal tract against the control (P < 0.001). Depending on the type of endoxylanase, a pool of AX with distinct physicochemical properties was created. The B. subtilis endoxylanase created a large pool of soluble AX in the ileum, thereby increasing ileal viscosity compared to broilers fed an endoxylanase from N. flexuosa (P < 0.001). The N. flexuosa endoxylanase mainly triggered caecal AX fermentation in young broilers, by delivering easily fermentable AX substrates with a low degree of polymerisation (P = 0.03). The effects were particularly present in young broilers (d 11). From this study, it is clear that the type and dose of endoxylanase added to wheat-soy based diets determine the nature of AX substrates formed. These, in turn, affect the intestinal viscosity and the interplay between the dietary AX compounds and microbiota, hence dictating AX digestion at young broiler ages and performance outcomes towards slaughter age.

Hydrothermal Treatments Cause Wheat Gluten-Derived Peptides to Form Amyloid-like Fibrils.

Formation of amyloid fibrils (i.e., protein structures containing a compact core of ordered ß-sheet structures) from food proteins can improve their techno-functional properties. Wheat gluten is the most consumed cereal protein by humans and extensively present in food and feed systems. Hydrolysis of wheat gluten increases the solubility of its proteins and brings new opportunities for value creation. In this study, the formation of amyloid-like fibrils (ALFs) from wheat gluten peptides (WGPs) under food relevant processing conditions was investigated. Different hydrothermal treatments were tested to maximize the formation of straight ALFs from WGPs. Thioflavin T (ThT) fluorescence measurements and transmission electron microscopy (TEM) were performed to study the extent of fibrillation and the morphology of the fibrils, respectively. First, the formation of fibrils by heating solutions of tryptic WGPs [degrees of hydrolysis 2.0% (DH 2) or 6.0% (DH 6)] was optimized using a response surface design. WGP solutions were incubated at different pH values, times, and temperatures. DH 6 WGPs had a higher propensity for fibrillation than did DH 2 WGPs. Heating DH 6 WGPs at 2.0% (w/v) for 38 h at 85 °C and pH 7.0 resulted in optimal fibrillation. Second, trypsin, chymotrypsin, thermolysin, papain, and proteinase K were used to produce different DH 6 WGPs. After enzyme inactivation and subsequent heating at optimal fibrillation conditions, chymotrypsin and proteinase K DH 6 WGPs produced small worm-like fibrils, whereas fibrils prepared from trypsin DH 6 WGPs were long and straight. The surface hydrophobicity of the peptides was key for fibrillation. Third, peptides from the wheat gluten components gliadin and glutenin fractions formed smaller and worm-like fibrils than did WGPs. Thus, the peptides of both gluten protein fractions jointly contribute to gluten fibrillation.

Solid-Phase PCR-Amplified DNAzyme Activity for Real-Time FO-SPR Detection of the MCR-2 Gene.

The polymerase chain reaction (PCR) has been the gold standard molecular analysis technique for decades and has seen quite some evolution in terms of reaction components, methodology, and readout mechanisms. Nucleic acid enzymes (NAzymes) have been used to further exploit the applications of PCR, but so far the work was limited to the colorimetric G-quadruplex or fluorescent substrate cleaving NAzymes. In this study, a solid-phase, fiber optic surface plasmon resonance (FO-SPR) technique is presented as an alternative readout for PCR utilizing NAzymes. First, the surface cleavage activity of DNAzyme-extended amplicons (DNAzyme-amps) is established, followed by optimization of the PCR conditions, which are required for compatibility with the FO-SPR system. Next, by integrating the complement of a 10-23 DNAzyme into the primer pair, PCR-amplified DNAzyme-amps were generated, tested, and validated on qPCR for the detection of the antimicrobial resistance gene MCR-2. Once validated, this primer concept was developed as a one-step assay, driven by PCR-amplified DNAzymes, for FO-SPR-based sensitive and specific detection. Using gold nanoparticle labeled RNA-DNA hybrid strands as substrate for the DNAzyme, PCR-amplified DNAzyme-amps generated in the presence of MCR-2 gene were monitored in real-time, which resulted in an experimental limit of detection of 4 × 105 copy numbers or 6.6 fM. In addition, the DNAzyme-based FO-PCR assay was able to discriminate between the MCR-1 and MCR-2 genes, to further prove the specificity of this assay. Henceforth, this DNAzyme-based fiber optic PCR assay provides a universally applicable, real-time system for the detection of virtually any target NA, in a specific and sensitive manner.

The impact of wheat (Triticum aestivum L.) bran on wheat starch gelatinization: A differential scanning calorimetry study.

The effect of wheat bran on starch gelatinization temperature was investigated. Dynamic water vapour sorption and water retention capacity experiments showed that bran bound up to 3 times more water than starch. However, examining starch gelatinization in starch-bran-water mixtures with differential scanning calorimetry showed that the effect of substituting starch by bran differed from that of moving into a regime of limiting water. Modelling the effect of the mixture composition on starch gelatinization behavior indicated that the onset (To) and peak (Tp) gelatinization temperatures were positively impacted by the bran concentration in water. The conclusion temperature (Tc) was negatively affected by the water content. Fractionation experiments demonstrated that the increased To and Tp were mainly caused by the extractable wheat bran components, such as potassium and phosphorus, which decrease the plasticization capacity of the solvent. The mechanism behind our observations was explained with the side-chain liquid-crystalline polymeric model for starch.

Revealing the main factors and two-way interactions contributing to food discolouration caused by iron-catechol complexation.

Fortification of food with iron is considered to be an effective approach to counter the global health problem caused by iron deficiency. However, reactivity of iron with the catechol moiety of food phenolics leads to discolouration and impairs bioavailability. In this study, we investigated the interplay between intrinsic and extrinsic factors on food discolouration caused by iron-catechol complexation. To this end, a three-level fractional factorial design was implemented. Absorbance spectra were analysed using statistical methods, including PCA, HCA, and ANOVA. Furthermore, a direct link between absorbance spectra and stoichiometry of the iron-catechol complexes was confirmed by ESI-Q-TOF-MS. All statistical methods confirm that the main effects affecting discolouration were type of iron salt, pH, and temperature. Additionally, several two-way interactions, such as type of iron salt × pH, pH × temperature, and type of iron salt × concentration significantly affected iron-catechol complexation. Our findings provide insight into iron-phenolic complexation-mediated discolouration, and facilitate the design of iron-fortified foods.

Study into the effect of microfluidisation processing parameters on the physicochemical properties of wheat (Triticum aestivum L.) bran.

The physicochemical properties of wheat bran have an effect on its technofunctional and nutritional profile. The possibility to induce physicochemical modifications in wheat bran using microfluidisation was investigated. An I-optimal experimental design was used to investigate the effect of microfluidisation processing parameters (pressure, number of passes, bran concentration and initial particle size) on important properties of wheat bran (particle size, microstructure, chemical composition, water retention capacity (WRC), extractability, viscosity and sedimentation). With the parameters used in this study, microfluidisation reduced wheat bran median particle size to 14.8 µm and disintegrated starch granules from the attached endosperm. This coincided with an increased extractability of starch and arabinoxylan. While the initial particle size was of minor importance, a higher pressure, larger number of passes and lower bran concentration during microfluidisation resulted in a smaller particle size, higher WRC and extractability, and an increased viscosity and stability in a 2% wheat bran suspension.

Impact of Nannochloropsis sp. dosage form on the oxidative stability of n-3 LC-PUFA enriched tomato purees.

Microalgae are a sustainable alternative source of n-3 LC-PUFA that can be incorporated into the food chain either via the incorporation of the (intact or disrupted) biomass or by the incorporation of the oil extracted from the biomass. However, the impact of the dosage form on the enrichment of food products with n-3 LC-PUFA and their oxidative stability has never been described before. This study aims to contribute more insight on the impact of the dosage form of the photoautotrophic microalga Nannochloropsis in enriched tomato puree. Three different dosage forms of Nannochloropsis were compared to commercial fish oil and analyzed for their amount of n-3 LC-PUFA, lipid oxidation products, antioxidants and free fatty acids. Tomato purees supplemented with dosage forms derived from Nannochloropsis showed higher oxidative stability than those supplemented with commercial fish oil. The highest oxidative stability was observed for purees supplemented with Nannochloropsis biomass irrespective of whether it was pre-disrupted.

Ultra-fast, sensitive and quantitative on-chip detection of group B streptococci in clinical samples.

PCR enables sensitive and specific detection of infectious disease agents, but application in point-of-care diagnostic testing remains scarce. A compact tool that runs PCR assays in less than a few minutes and that relies on mass-producible, disposable reactors could revolutionize while-you-wait molecular testing. We here exploit well-established semiconductor manufacturing processes to produce silicon ultra-fast quantitative PCR (UF-qPCR) chips that can run PCR protocols with limited assay optimization. A total of 110 clinical samples were analyzed for the detection of group B streptococci using both a validated benchtop and an on-chip qPCR assay. For the on-chip assay, the total reaction time was reduced after optimization to less than 5 min. The standard curve, spanning a concentration range of 5 log units, yielded a PCR efficiency of 94%. The sensitivity obtained was 96% (96/100; CI: 90-98%) and the specificity 70% (7/10; CI: 40-90%). We show that if melting analyses would be integrated, the obtained sensitivity would drop slightly to 93% (CI: 86-96%), while the specificity would increase to 100% (CI: 72% - 100%). In comparison to the benchtop reference qPCR assay performed on a LightCycler©96, the on-chip assay demonstrated a highly significant qualitative (Spearman's rank correlation) and quantitative (linear regression) correlation. Using a mass-producible qPCR chip and limited assay optimization, we were able to develop a validated qPCR protocol that can be carried out in less than five minutes. The analytical performance of the microchip-based UF-qPCR system was shown to match that of a benchtop assay. This is the first report to provide UF-qPCR validation using clinical samples. We demonstrate that qPCR-based while-you-wait testing is feasible without jeopardizing assay performance.

The impact of steeping, germination and hydrothermal processing of wheat (Triticum aestivum L.) grains on phytate hydrolysis and the distribution, speciation and bio-accessibility of iron and zinc elements.

Chelation of iron and zinc in wheat as phytates lowers their bio-accessibility. Steeping and germination (15 °C, 120 h) lowered phytate content from 0.96% to only 0.81% of initial dry matter. A multifactorial experiment in which (steeped/germinated) wheat was subjected to different time (2-24 h), temperature (20-80 °C) and pH (2.0-8.0) conditions showed that hydrothermal processing of germinated (15 °C, 120 h) wheat at 50 °C and pH 3.8 for 24 h reduced phytate content by 95%. X-ray absorption near-edge structure imaging showed that it indeed abolished chelation of iron to phytate. It also proved that iron was oxidized during steeping, germination and hydrothermal processing. It was further shown that zinc and iron bio-accessibility were respectively 3 and 5% in wheat and 27 and 37% in hydrothermally processed wheat. Thus, hydrothermal processing of (germinated) wheat paves the way for increasing elemental bio-accessibility in whole grain-based products.

Assessing the relative importance of health and conformation traits in the cavalier king Charles spaniel.

BACKGROUND: The selection of a future breeding dog is a complicated task, in which disease characteristics and different traits have to be combined and weighed against one another. Truncation selection, that is the exclusion of affected animals, may be very inefficient when selecting on a large number of traits, and may result in a reduction of the genetic diversity in a population or breed. Selection could be facilitated by the use of a selection index that combines multiple traits or breeding values into one score. This however requires a consideration of their relative value according to their economic weight, which is difficult to express in monetary units for health traits. The use of a choice experiment to derive non-market values might be a solution to this problem. This is a pilot study to assess the potential use of choice experiments to ascertain the public preference and relative importance attached to health- and conformation traits in the selection of a Cavalier King Charles spaniel. The focus was on two prevalent disorders, mitral valve disease and syringomyelia, and on several important conformation traits such as muzzle length and eye shape. Based on available prior information, a Bayesian D-optimal design approach was used to develop a choice experiment and the resulting choice sets. RESULTS: Every participant (breeder or owner) in the choice experiment was presented with a total of 17 choice sets, in which at most four traits could vary to reduce the cognitive burden. A total of 114 respondents participated in the choice experiment and results showed that respondents (breeders/owners) current attitudes were directed towards health (syringomyelia and mitral valve disease), followed by eye shape and level of inbreeding. CONCLUSIONS: This approach identifies the value breeders and owners attach to certain traits in the breeding objective. The resulting relative weights, represented as the logworths obtained from the choice experiment, could be an alternative to economic weights. They could be implemented as a weight when breeding values are available, but more study on this topic will be necessary. A challenge in this approach is to scale up the experiment with additional traits. Moreover, for other traits, the genetic parameters and correlations should be known first, in order to include them in the health selection index as well.

Optical Manipulation of Single Magnetic Beads in a Microwell Array on a Digital Microfluidic Chip.

The detection of single molecules in magnetic microbead microwell array formats revolutionized the development of digital bioassays. However, retrieval of individual magnetic beads from these arrays has not been realized until now despite having great potential for studying captured targets at the individual level. In this paper, optical tweezers were implemented on a digital microfluidic platform for accurate manipulation of single magnetic beads seeded in a microwell array. Successful optical trapping of magnetic beads was found to be dependent on Brownian motion of the beads, suggesting a 99% chance of trapping a vibrating bead. A tailor-made experimental design was used to screen the effect of bead type, ionic buffer strength, surfactant type, and concentration on the Brownian activity of beads in microwells. With the optimal conditions, the manipulation of magnetic beads was demonstrated by their trapping, retrieving, transporting, and repositioning to a desired microwell on the array. The presented platform combines the strengths of digital microfluidics, digital bioassays, and optical tweezers, resulting in a powerful dynamic microwell array system for single molecule and single cell studies.

Augmented design and analysis of computer experiments: a novel tolerance embedded global optimization approach applied to SWIR hyperspectral illumination design.

A novel meta-heuristic approach for minimizing nonlinear constrained problems is proposed, which offers tolerance information during the search for the global optimum. The method is based on the concept of design and analysis of computer experiments combined with a novel two phase design augmentation (DACEDA), which models the entire merit space using a Gaussian process, with iteratively increased resolution around the optimum. The algorithm is introduced through a series of cases studies with increasing complexity for optimizing uniformity of a short-wave infrared (SWIR) hyperspectral imaging (HSI) illumination system (IS). The method is first demonstrated for a two-dimensional problem consisting of the positioning of analytical isotropic point sources. The method is further applied to two-dimensional (2D) and five-dimensional (5D) SWIR HSI IS versions using close- and far-field measured source models applied within the non-sequential ray-tracing software FRED, including inherent stochastic noise. The proposed method is compared to other heuristic approaches such as simplex and simulated annealing (SA). It is shown that DACEDA converges towards a minimum with 1 % improvement compared to simplex and SA, and more importantly requiring only half the number of simulations. Finally, a concurrent tolerance analysis is done within DACEDA for to the five-dimensional case such that further simulations are not required.

Relative importance and interactions of furan precursors in sterilised, vegetable-based food systems.

Mitigation strategies aimed at an intervention in the reaction pathways for furan formation (e.g., by adjusting precursor concentrations) might offer an additional route for furan reduction in sterilised, vegetable-based foods, without adverse effects on other food safety or quality attributes. As a first step towards product reformulation, the aim of the present study was to determine the relative importance and interactions of possible furan precursors in these types of foods. Based on an I-optimal experimental design, potato purée (naturally low in furan precursors) was spiked with known amounts of sugars, ascorbic acid, olive oil and ß-carotene, and subjected to a thermal sterilisation. Significant correlations were observed between furan concentrations after thermal treatment and starting concentrations of ascorbic acid and monosaccharides (i.e., fructose and glucose). Ascorbic acid had a clear furan-reducing effect as an antioxidant by protecting (polyunsaturated) fatty acids against oxidative degradation. Fructose and glucose were the main precursors, which can most probably be attributed to their high, but realistic, concentrations in the product. The contributions of fatty acids and ß-carotene were strongly dependent on redox interactions with other food constituents. In the same potato purées, only low concentrations (0-2 ng g(-1) purée) of 2-methylfuran were detected, indicating that the direct importance of the spiked food constituents as a precursor for methylfuran formation was rather small. Based on the results of this study, reducing the amount of monosaccharides or adjusting the redox conditions of the matrix are suggested as two possible approaches for furan mitigation on the product side.

Public preferences for prioritizing preventive and curative health care interventions: a discrete choice experiment.

BACKGROUND: Setting fair health care priorities counts among the most difficult ethical challenges our societies are facing. OBJECTIVE: To elicit through a discrete choice experiment the Belgian adult population's (18-75 years; N = 750) preferences for prioritizing health care and investigate whether these preferences are different for prevention versus cure. METHODS: We used a Bayesian D-efficient design with partial profiles, which enables considering a large number of attributes and interaction effects. We included the following attributes: 1) type of intervention (cure vs. prevention), 2) effectiveness, 3) risk of adverse effects, 4) severity of illness, 5) link between the illness and patient's health-related lifestyle, 6) time span between intervention and effect, and 7) patient's age group. RESULTS: All attributes were statistically significant contributors to the social value of a health care program, with patient's lifestyle and age being the most influential ones. Interaction effects were found, showing that prevention was preferred to cure for disease in young adults, as well as for severe and lethal disease in people of any age. However, substantial differences were found in the preferences of respondents from different age groups, with different lifestyles and different health states. CONCLUSIONS: Our study suggests that according to the Belgian public, contextual factors of health gains such as patient's age and health-related lifestyle should be considered in priority setting decisions. The studies, however, revealed substantial disagreement in opinion between different population subgroups.

Historical land use change has lowered terrestrial silica mobilization.

Continental export of Si to the coastal zone is closely linked to the ocean carbon sink and to the dynamics of phytoplankton blooms in coastal ecosystems. Presently, however, the impact of human cultivation of the landscape on terrestrial Si fluxes remains unquantified and is not incorporated in models for terrestrial Si mobilization. In this paper, we show that land use is the most important controlling factor of Si mobilization in temperate European watersheds, with sustained cultivation (>250 years) of formerly forested areas leading to a twofold to threefold decrease in baseflow delivery of Si. This is a breakthrough in our understanding of the biogeochemical Si cycle: it shows that human cultivation of the landscape should be recognized as an important controlling factor of terrestrial Si fluxes.