Xeno-Free 3D Bioprinted Liver Model for Hepatotoxicity Assessment.

Three-dimensional (3D) bioprinting is one of the most promising methodologies that are currently in development for the replacement of animal experiments. Bioprinting and most alternative technologies rely on animal-derived materials, which compromises the intent of animal welfare and results in the generation of chimeric systems of limited value. The current study therefore presents the first bioprinted liver model that is entirely void of animal-derived constituents. Initially, HuH-7 cells underwent adaptation to a chemically defined medium (CDM). The adapted cells exhibited high survival rates (85-92%) after cryopreservation in chemically defined freezing media, comparable to those preserved in standard medium (86-92%). Xeno-free bioink for 3D bioprinting yielded liver models with high relative cell viability (97-101%), akin to a Matrigel-based liver model (83-102%) after 15 days of culture. The established xeno-free model was used for toxicity testing of a marine biotoxin, okadaic acid (OA). In 2D culture, OA toxicity was virtually identical for cells cultured under standard conditions and in CDM. In the xeno-free bioprinted liver model, 3-fold higher concentrations of OA than in the respective monolayer culture were needed to induce cytotoxicity. In conclusion, this study describes for the first time the development of a xeno-free 3D bioprinted liver model and its applicability for research purposes.

Potential of Modification of Techno-Functional Properties and Structural Characteristics of Citrus, Apple, Oat, and Pea Dietary Fiber by High-Intensity Ultrasound.

Plant fibers are rich in dietary fiber and micronutrients but often exhibit poor functionality. Ultrasonication can affect the particle size of plant fiber, thereby influencing other techno-functional properties. Therefore, this study aimed to investigate the effects of high-intensity ultrasound on citrus, apple, oat, and pea fiber. Initially, solutions containing 1 wt% of plant fiber were homogenized using ultrasonication (amplitude 116 µm, t = 150 s, energy density = 225 kJ/L, P¯ = 325 W). Due to cavitation effects induced by ultrasound, differences in particle size and a shift in the ratio of insoluble and alcohol-insoluble fractions for dietary fiber were observed. Additionally, viscosities for citrus and apple fiber increased from 1.4 Pa·s to 84.4 Pa·s and from 1.34 Pa·s to 31.7 Pa·s, respectively, at shear rates of 100 1s. This was attributed to observed differences in the microstructure. Freeze-dried samples of purified citrus and apple fiber revealed thin and nearly transparent layers, possibly contributing to enhanced water binding capacity and, therefore, increased viscosity. Water binding capacity for citrus fiber increased from 18.2 g/g to 41.8 g/g, and a 40% increase was observed for apple fiber. Finally, ultrasound demonstrated itself be an effective technology for modifying the techno-functional properties of plant fiber, such as water binding capacity.

Influence of High-Intensity Ultrasound on Characteristics and Bioaccessibility of Pea Protein in Fiber-Enriched Suspensions.

Pea protein is of high interest for the food industry owing to its low allergenicity and high nutritional value. However, it often exhibits poor functionality, such as low solubility. The presence of dietary fiber in food products is beneficial for human health but may decrease the bioaccessibility of nutrients. Ultrasound, as a promising green technology, may influence properties of fibers and proteins and, thus, bioaccessibility. Therefore, this study investigated the effects of high-intensity ultrasound on the characteristics and protein bioaccessibility of protein-fiber suspensions. Suspensions containing different fiber compounds (1 wt.%) and pea protein (5 wt.%) were homogenized using high-intensity ultrasound (amplitude 116 µm, t = 150 s, energy density = 225 kJ/L, P¯ = 325 W). Owing to sonication-induced cavitation, the dispersibility of the protein was enhanced, and the viscosity of solutions containing citrus or apple fiber was increased. FE-SEM revealed the formation of different fiber-protein networks during sonication. Even if viscosity is known to have an impact on the bioaccessibility of nutrients, no restrictions on the digestibility of protein were detected during an in vitro digestion. Thus, protein uptake is probably not affected, and ultrasound can be used to modify the technofunctionality of fibers and proteins without any nutritional disadvantages.

Towards a Better Understanding of Texturization during High-Moisture Extrusion (HME)-Part II: Characterization of Thermophysical Properties of High-Moisture Meat Analogues.

It is crucial to determine the thermophysical properties of high-moisture extruded samples (HMESs) to properly understand the texturization process of high-moisture extrusion (HME), especially when the primary objective is the production of high-moisture meat analogues (HMMAs). Therefore, the study's aim was to determine thermophysical properties of high-moisture extruded samples made from soy protein concentrate (SPC ALPHA® 8 IP). Thermophysical properties such as the specific heat capacity and the apparent density were experimentally determined and further investigated to obtain simple prediction models. These models were compared to non-HME-based literature models, which were derived from high-moisture foods, such as soy-based and meat products (including fish). Furthermore, thermal conductivity and thermal diffusivity were calculated based on generic equations and literature models and showed a significant mutual influence. The combination of the experimental data and the applied simple prediction models resulted in a satisfying mathematical description of the thermophysical properties of the HME samples. The application of data-driven thermophysical property models could contribute to understanding the texturization effect during HME. Further, the gained knowledge could be applied for further understanding in related research, e.g., with numerical simulation studies of the HME process.

Towards a Better Understanding of Texturization during High-Moisture Extrusion (HME)-Part I: Modeling the Texturability of Plant-Based Proteins.

This study focused on predicting high-moisture texturization of plant-based proteins (soy protein concentrate (SPC), soy protein isolate (SPI), pea protein isolate (PPI)) at different water contents (57.5%, 60%, 65%, 70%, and 72.5% (w/w db)) to optimize and guarantee the production of high-moisture meat analogs (HMMA). Therefore, high-moisture extrusion (HME) experiments were performed, and the texture of the obtained high-moisture extruded samples (HMES) was sensory evaluated and categorized into poorly-textured, textured, or well-textured. In parallel, data on heat capacity (cp) and phase transition behavior of the plant-based proteins were determined using differential scanning calorimetry (DSC). Based on the DSC data, a model for predicting cp of hydrated, but not extruded, plant-based proteins was developed. Furthermore, based on the aforementioned model for predicting cp and DSC data on phase transition behavior of the plant-based proteins in combination with conducted HME trials and the mentioned model for predicting cp, a texturization indicator was developed, which could be used to calculate the minimum threshold temperature required to texturize plant-based proteins during HME. The outcome of this study could help to minimize the resources of expensive extrusion trials in the industry to produce HMMA with defined textures.

Differentiation of sea buckthorn syrups processed by high pressure, pulsed electric fields, ohmic heating, and thermal pasteurization based on quality evaluation and chemical fingerprinting.

Introduction: Impact of processing on product characteristics, sustainability, traceability, authenticity, and public health along the food chain becomes more and more important not only to the producer but also to the customer and the trust of a consumer toward a brand. In recent years, the number of juices and smoothies containing so called super foods or fruits, which have been "gently pasteurized," has increased significantly. However, the term "gentle pasteurization" related to the application of emerging preservation technologies such as pulsed electric fields (PEF), high pressure processing (HPP) or ohmic heating (OH) is not clearly defined. Methods: Therefore, the presented study investigated the influence of PEF, HPP, OH, and thermal treatment on quality characteristics and microbial safety of sea buckthorn syrup. Syrups from two different varieties were investigated under the following conditions HPP (600 MPa 4-8 min), OH (83°C and 90°C), PEF (29.5 kV/cm, 6 µs, 100 Hz), and thermal (88°C, hot filling). Analyses to test the influence on quality parameters like ascorbic acid (AA), flavonoids, carotenoids, tocopherols, antioxidant activity; metabolomical/chemical profiling (fingerprinting) via U-HPLC-HRMS/MS (here especially flavonoids and fatty acids); sensory evaluation, as well as microbial stability including storage, were conducted. Results and discussion: Independent from the treatment, the samples were stable over 8 weeks of storage at 4°C. The influence on the nutrient content [Ascorbic acid (AA), total antioxidant activity (TAA), total phenolic compounds (TPC), tocopherols (Vit E)] was similar for all tested technologies. Employing statistical evaluation Principal Component Analysis (PCA) a clear clustering based on the processing technologies was observed. Flavonoids as well as fatty acids were significantly impacted by the type of used preservation technology. This was obvious during the storage time of PEF and HPP syrups, where enzyme activity was still active. The color as well as taste of the syrups were found to be more fresh-like for the HPP treated samples.

A new approach for calculating microalgae culture growth based on an inhibitory effect of the surrounding biomass.

Ever since the potential of algae in biotechnology was recognized, models describing the growth of algae inside photobioreactors have been proposed. These models are the basis for the optimization of process conditions and reactor designs. Over the last few decades, models became more and more elaborate with the increase of computational capacity. Thus far, these models have been based on light attenuation due to the absorption and scattering effects of the biomass. This manuscript presents a new way of predicting the apparent growth inside photobioreactors using simple models for enzymatic kinetics to describe the reaction between photons and the photosynthetic unit. The proposed model utilizes an inhibition kinetic formula based on the surrounding biomass to describe the average growth rate of a culture, which is determined by the local light intensities inside the reactor. The result is a mixed-inhibition scheme with multiple inhibition sites. The parameters of the new kinetic equation are replaced by empirical regression functions to correlate their dependency on incident light intensity and reactor size. The calibrations of the parameters and the regression functions are based on the numerical solutions of the growth rate computed with a classical Type II model. As a final verification, we apply the new equation in predicting the growth behavior of three phototrophic organisms in reactors of three different sizes.

Structure and adsorption behavior of high hydrostatic pressure-treated ß-lactoglobulin.

HYPOTHESIS: High hydrostatic pressure treatment causes structural changes in interfacial-active ß-lactoglobulin (ß-lg). We hypothesized that the pressure-induced structural changes affect the intra- and intermolecular interactions which determine the interfacial activity of ß-lg. The conducted experimental and numerical investigations could contribute to the mechanistic understanding of the adsorption behavior of proteins in food-related emulsions. EXPERIMENTS: We treated ß-lg in water at pH 7 with high hydrostatic pressures up to 600 MPa for 10 min at 20 °C. The secondary structure was characterized with Fourier-transform infrared spectroscopy (FTIR) and circular dichroism (CD), the surface hydrophobicity and charge with fluorescence-spectroscopy and ζ-potential, and the quaternary structure with membrane-osmometry, analytical ultracentrifugation (AUC) and mass spectrometry (MS). Experimental analyses were supported through molecular dynamic (MD) simulations. The adsorption behavior was investigated with pendant drop analysis. FINDINGS: MD simulation revealed a pressure-induced molten globule state of ß-lg, confirmed by an unfolding of ß-sheets with FTIR, a stabilization of α-helices with CD and loss in tertiary structure induced by an increase in surface hydrophobicity. Membrane-osmometry, AUC and MS indicated the formation of non-covalently linked dimers that migrated slower through the water phase, adsorbed more quickly due to hydrophobic interactions with the oil, and lowered the interfacial tension more strongly than reference ß-lg.

Concentrating Model Solutions and Fruit Juices Using CO2 Hydrate Technology and Its Quantitative Effect on Phenols, Carotenoids, Vitamin C and Betanin.

Fruits have an important economic impact in the context of plant-based food production. The consumption of fruit juices, mostly produced from concentrates, is particularly noteworthy. Conventional concentration methods do not always enable a sustainable and gentle concentration. The innovative gas hydrate technology addresses this point with its energy-saving, gentle character, and high concentration potential. In this study, the concentration of fruit juices and model solutions using CO2 hydrate technology was investigated. To find a suitable operating point for hydrate formation in the used bubble column, the hydrate formation in a water-sucrose model solution was evaluated at different pressure and temperature combinations (1, 3, 5 °C and 32.5, 37.5, 40 bar). The degrees of concentration indicate that the bubble column reactor operates best at 37.5 bar and 3 °C. To investigate the gentle processing character of the hydrate technology, its quantitative effects on vitamin C, betanin, polyphenols, and carotenoids were analyzed in the produced concentrates and hydrates via HPLC and UV/VIS spectrophotometry. The results for fruit juices and model solutions imply that all examined substances are accumulated in the concentrate, while only small amounts remain in the hydrate. These amounts can be related to an inefficient separation process.

Gentle Sterilization of Carrot-Based Purees by High-Pressure Thermal Sterilization and Ohmic Heating and Influence on Food Processing Contaminants and Quality Attributes.

Pressure-enhanced sterilization (PES) and ohmic heating (OH) are two emerging sterilization techniques, currently lacking implementation in the food industry. However, both technologies offer significant benefits in terms of spore inactivation using reduced thermal intensity in food products, as well as minimized effects on sensory and nutritional profiles. In this study, PES and OH were tested based on possible food safety process windows in comparison to thermal retorting, to optimize the food quality of carrot-based purees. The following parameters related to food quality were tested: texture, carotenoid content, color, and detectable amount of food processing contaminants (FPC) formed. Application of the innovative sterilization techniques resulted in a better retention of color, texture, and carotenoids (for PES) as well as a reduced formation of food processing contaminants. Importantly, a significant reduction in the formation of furan and its derivates was observed, compared to the retorted samples. Hence, both sterilization technologies showed promising results in the mitigation of potential toxic processing contaminants and retention of quality attributes.

Development of a Continuous Pulsed Electric Field (PEF) Vortex-Flow Chamber for Improved Treatment Homogeneity Based on Hydrodynamic Optimization.

Pulsed electric fields (PEF) treatment is an effective process for preservation of liquid products in food and biotechnology at reduced temperatures, by causing electroporation. It may contribute to increase retention of heat-labile constituents with similar or enhanced levels of microbial inactivation, compared to thermal processes. However, especially continuous PEF treatments suffer from inhomogeneous treatment conditions. Typically, electric field intensities are highest at the inner wall of the chamber, where the flow velocity of the treated product is lowest. Therefore, inhomogeneities of the electric field within the treatment chamber and associated inhomogeneous temperature fields emerge. For this reason, a specific treatment chamber was designed to obtain more homogeneous flow properties inside the treatment chamber and to reduce local temperature peaks, therefore increasing treatment homogeneity. This was accomplished by a divided inlet into the chamber, consequently generating a swirling flow (vortex). The influence of inlet angles on treatment homogeneity was studied (final values: radial angle α = 61°; axial angle ß = 98°), using computational fluid dynamics (CFD). For the final design, the vorticity, i.e., the intensity of the fluid rotation, was the lowest of the investigated values in the first treatment zone (1002.55 1/s), but could be maintained for the longest distance, therefore providing an increased mixing and most homogeneous treatment conditions. The new design was experimentally compared to a conventional co-linear setup, taking into account inactivation efficacy of Microbacterium lacticum as well as retention of heat-sensitive alkaline phosphatase (ALP). Results showed an increase in M. lacticum inactivation (maximum Δlog of 1.8 at pH 7 and 1.1 at pH 4) by the vortex configuration and more homogeneous treatment conditions, as visible by the simulated temperature fields. Therefore, the new setup can contribute to optimize PEF treatment conditions and to further extend PEF applications to currently challenging products.

Kinetic Modeling and Numerical Simulation as Tools to Scale Microalgae Cell Membrane Permeabilization by Means of Pulsed Electric Fields (PEF) From Lab to Pilot Plants.

Pulsed Electric Fields (PEF) is a promising technology for the gentle and energy efficient disruption of microalgae cells such as Chlorella vulgaris. The technology is based on the exposure of cells to a high voltage electric field, which causes the permeabilization of the cell membrane. Due to the dependency of the effective treatment conditions on the specific design of the treatment chamber, it is difficult to compare data obtained in different chambers or at different scales, e.g., lab or pilot scale. This problem can be overcome by the help of numerical simulation since it enables the accessibility to the local treatment conditions (electric field strength, temperature, flow field) inside a treatment chamber. To date, no kinetic models for the cell membrane permeabilization of microalgae are available what makes it difficult to decide if and in what extent local treatment conditions have an impact on the permeabilization. Therefore, a kinetic model for the perforation of microalgae cells of the species Chlorella vulgaris was developed in the present work. The model describes the fraction of perforated cells as a function of the electric field strength, the temperature and the treatment time by using data which were obtained in a milliliter scale batchwise treatment chamber. Thereafter, the model was implemented in a CFD simulation of a pilot-scale continuous treatment chamber with colinear electrode arrangement. The numerical results were compared to experimental measurements of cell permeabilization in a similar continuous treatment chamber. The predicted values and the experimental data agree reasonably well what demonstrates the validity of the proposed model. Therefore, it can be applied to any possible treatment chamber geometry and can be used as a tool for scaling cell permeabilization of microalgae by means of PEF from lab to pilot scale. The present work provides the first contribution showing the applicability of kinetic modeling and numerical simulation for designing PEF processes for the purpose of biorefining microalgae biomass. This can help to develop new processes and to reduce the costs for the development of new treatment chamber designs.

Matrix- and Technology-Dependent Stability and Bioaccessibility of Strawberry Anthocyanins during Storage.

Anthocyanins are often associated with health benefits. They readily degrade during processing and storage but are also dependent on the matrix conditions. This study investigated how strawberry anthocyanins are affected by preservation technologies and a relatively protein-rich kale juice addition during storage. A strawberry-kale mix was compared to a strawberry-water mix (1:2 wt; pH 4), untreated, thermally, pulsed electric fields (PEF) and high-pressure processing (HPP) treated, and evaluated for anthocyanin stability and bioaccessibility during refrigerated storage. The degradation of strawberry anthocyanins during storage followed first-order kinetics and was dependent on the juice system, preservation technology and anthocyanin structure. Generally, the degradation rate was higher for the strawberry-kale mix compared to the strawberry-water mix. The untreated sample showed the highest degradation rate, followed by HPP, PEF and, then thermal. The relative anthocyanin bioaccessibility after gastric digestion was 10% higher for the thermally and PEF treated samples. Anthocyanin bioaccessibility after intestinal digestion was low due to instability at a neutral pH, especially for the strawberry-kale mix, and after thermal treatment. The storage period did not influence the relative bioaccessibility; yet, the absolute content of bioaccessible anthocyanins was decreased after storage. This research further presents that processing and formulation strongly affect the stability and bioaccessibility of anthocyanins during storage.

Comparative analysis of bioactive phenolic compounds composition from 26 medicinal plants.

Bioactive phenolic compounds are powerful antioxidants in traditionally used medicinal and industrial crop plants and have attracted increased interest in the last years in their application and role in non-destructive methodology for pre-screening analysis of some stress factors. In this study the qualitative target was linked with future possible applications of received data for improving non-destructive methodology as well as for improving existing knowledge regarding antioxidant content in some plant species. Comparative analysis of total phenolics, flavonoid contents, phenolic acid composition, and antioxidant activity in known east central Europe medicinal and industrial crop plants of 26 species of families Asteraceae, Rosaceae and Lamiaceae was done. Among the investigated leaf extracts the highest total phenolic, total flavonoid contents and antioxidant activity have been seen for Stachys byzantine L. (Lamiaceae), Calendula officinalis L. (Asteraceae) and for Potentilla recta L. (Rosaceae). The highest syringic acid content has been found in the leaf extracts of plant family Asteraceae - in the range from 0.782 to 5.078 mg g-1 DW. The representative's family Rosaceae has a higher content of p-anisic acid in the range 0.334-3.442 mg g-1DW compared to the leaf extracts of families Lamiaceae and Asteraceae. The comparative study showed significant differences of content of phenolic acids in the leaf extracts of different representative's families Rosaceae, Asteraceae and Lamiaceae. We suggest that the presence of some phenolic acids can be used as a possible marker for family botanical specifications of representative families Asteraceae and Rosaceae. It was supposed that some pharmacological effects can be connected with the analyzed data.

Soil Biodiversity Effects from Field to Fork.

Our knowledge of soil biodiversity in agriculture in general is currently increasing rapidly. However, almost all studies have stopped with the quantification of soil biodiversity effects on crops at harvest time, ignoring subsequent processes along the agrifood chain until food arrives on our plates. Here we develop a conceptual framework for the study of such postharvest effects. We present the main mechanisms (direct and indirect) via which soil biodiversity can influence crop quality aspects and give examples of how effects at harvest time may become attenuated through postharvest operations and how biodiversity may also affect some of these operations (i.e., storage) themselves. Future research with a broader focus has the potential to unveil how soil biodiversity may benefit from what ends up on our forks.

New lattice Boltzmann method for the simulation of three-dimensional radiation transfer in turbid media.

Based on the kinetic theory of photons, a new lattice Boltzmann method for the simulation of 3D radiation transport is presented. The method was successfully validated with Monte Carlo simulations of radiation transport in optical thick absorbing and non-absorbing turbid media containing either isotropic or anisotropic scatterers. Moreover, for the approximation of Mie-scattering, a new iterative algebraic approach for the discretization of the scattering phase function was developed, ensuring full conservation of energy and asymmetry after discretization. It was found that the main error sources of the method are caused by linearization and ray effects and suggestions for further improvement of the method are made.

Impact of surface structure and feed gas composition on Bacillus subtilis endospore inactivation during direct plasma treatment.

This study investigated the inactivation efficiency of cold atmospheric pressure plasma treatment on Bacillus subtilis endospores dependent on the used feed gas composition and on the surface, the endospores were attached on. Glass petri-dishes, glass beads, and peppercorns were inoculated with the same endospore density and treated with a radio frequency plasma jet. Generated reactive species were detected using optical emission spectroscopy. A quantitative polymerase chain reaction (qPCR) based ratio detection system was established to monitor the DNA damage during the plasma treatment. Argon + 0.135% vol. oxygen + 0.2% vol. nitrogen as feed gas emitted the highest amounts of UV-C photons and considerable amount of reactive oxygen and nitrogen species. Plasma generated with argon + 0.135% vol. oxygen was characterized by the highest emission of reactive oxygen species (ROS), whereas the UV-C emission was negligible. The use of pure argon showed a negligible emission of UV photons and atomic oxygen, however, the emission of vacuum (V)UV photons was assumed. Similar maximum inactivation results were achieved for the three feed gas compositions. The surface structure had a significant impact on the inactivation efficiency of the plasma treatment. The maximum inactivation achieved was between 2.4 and 2.8 log10 on glass petri-dishes and 3.9 to 4.6 log10 on glass beads. The treatment of peppercorns resulted in an inactivation lower than 1.0 log10. qPCR results showed a significant DNA damage for all gas compositions. Pure argon showed the highest results for the DNA damage ratio values, followed by argon + 0.135% vol. oxygen + 0.2% vol. nitrogen. In case of argon + 0.135% vol. oxygen the inactivation seems to be dominated by the action of ROS. These findings indicate the significant role of VUV and UV photons in the inactivation process of B. subtilis endospores.

Wavelet synthetic method for turbulent flow.

Based on the idea of random cascades on wavelet dyadic trees and the energy cascade model known as the wavelet p model, a series of velocity increments in two-dimensional space are constructed in different levels of scale. The dynamics is imposed on the generated scales by solving the Euler equation in the Lagrangian framework. A dissipation model is used in order to cover the shortage of the p model, which only predicts in inertial range. Wavelet reconstruction as well as the multiresolution analysis are then performed on each scales. As a result, a type of isotropic velocity field is created. The statistical properties show that the constructed velocity fields share many important features with real turbulence. The pertinence of this approach in the prediction of flow intermittency is also discussed.

Multifractal-cascade model for inertial and dissipation ranges based on the wavelet reconstruction method.

The discrete wavelet is introduced to construct the turbulent velocity fields. The simple binary cascade model p model is served as the inertial range model for velocity increments. The dissipation model, which follows Foias et al. [Phys. Fluids A 2, 464 (1990)] takes the form of exp(-gk). The length of inertial and dissipation ranges is computed according to the different construction levels. Based on the binary cascade theory and the proposed dissipation model, the Reynolds number regarding to the cascade process can be estimated. The dissipation rate calculated from the proposed model not only agrees with the existing experiment data, but also suggests that the dissipation rate is not an independent variable with respect to the Reynolds number.