Efficiency of aqueous oleosome extraction from capsicum seeds compared to classical oil extraction.

The extraction of oil from oilseeds in intact oleosomes is one of the suggested processes that could replace the extraction of oil by pressing and solvent extraction, being milder, environmentally less impactful and potentially more efficient in its use of resources. This study assesses the latter using an exergy assessment of oleosome extraction for food emulsions. The contribution of each part of the process to the overall impact was investigated. Based on current lab-scale data, oleosome extraction has nearly twice the exergy loss compared to the industrial process of oil extraction and industrial assembly of emulsions. The exergy losses of the lab-scale oleosome extraction are currently dominated by the chemical exergy associated with product loss during the separation of oleosomes from the rest of the biomass. This loss is expected to significantly decrease when upscaled to industrial scale. When substituted with industrial material efficiencies, the total exergy loss decreased to nearly a quarter of the original loss, representing oleosome extraction as a potentially more effective and environment-friendly option.

Bovine Serum Albumin Rejection by an Open Ultrafiltration Membrane: Characterization and Modeling.

The classic application of ultrafiltration (UF) is for the complete retention of proteins, and in that situation, the transport behavior is well established. More open membranes with fractional retention are used when separating different proteins. However, protein transport has not been well documented yet in the literature. The bovine serum albumin (∼69 kDa) observed rejection ranges from 0.65 to 1 using a 300 kDa molecular weight cut-off membrane at different pH, ionic strength, and pressure. We demonstrated that, especially with open UF, the transport of proteins through the membrane is dominated by advection, with insignificant diffusion effects (p value > 0.05). We showed that with open UF, retention is not only caused by size exclusion but also to a large extent by electrostatic interactions and oligomerization of the proteins. Mass transfer in the polarization layer was relatively independent of the pH and ionic strength. It was underestimated by common Sherwood relations due to a relatively large contribution of the reduction in the flow turbulence near the membrane by the removal of fluid through the membrane. We propose a model that allows relatively quick characterization of the rejection of proteins without prior knowledge of the pore sizes and charges based on just a limited set of experiments. Therefore, protein rejection with the open UF system can be targeted by tuning the processing conditions, which might be useful for designing protein fractionation processes.

Engineering artificial casein micelles for future food: Is casein phosphorylation necessary?

Industrial-scale production of recombinant proteins for food products may become economically feasible but correct post-translational modification of proteins by microbial expression systems remains a challenge. For efficient production of hybrid products from bovine casein and recombinant casein, it is therefore of interest to evaluate the necessity of casein post-translational phosphorylation for the preparation of hybrid casein micelles and study their rennet-induced coagulation. Our results show that dephosphorylated casein was hardly incorporated into artificial casein micelles but was capable of stabilising calcium phosphate nanoclusters with an increased size through adsorption on their surface. Thereby, dephosphorylated casein formed larger colloidal particles with a decreased hydration. Furthermore, the presence of increasing amounts of dephosphorylated casein resulted in increasingly poor rennet coagulation behaviour, where dephosphorylated casein disrupted the formation of a coherent gel network by native casein. These results emphasise that post-translational phosphorylation of casein is crucial for their assembly into micelles and thereby for the production of dairy products for which the casein micelle structure is a prerequisite, such as many cheese varieties and yoghurt. Therefore, phosphorylation of future recombinant casein is essential to allow its use in the production of animal-free dairy products.

Amyloid-like aggregation of recombinant ß-lactoglobulin at pH 3.5 and 7.0: Is disulfide bond removal the key to fibrillation?

Functional nanofibrils from globular proteins are usually formed by heating for several hours at pH 2.0, which induces acidic hydrolysis and consecutive self-association. The functional properties of these micro-metre-long anisotropic structures are promising for biodegradable biomaterials and food applications, but their stability at pH > 2.0 is low. The results presented here show that modified ß-lactoglobulin can also form nanofibrils by heating at neutral pH without prior acidic hydrolysis; the key is removing covalent disulfide bonds via precision fermentation. The aggregation behaviour of various recombinant ß-lactoglobulin variants was systemically studied at pH 3.5 and 7.0. The suppression of intra- and intermolecular disulfide bonds by eliminating one to three out of the five cysteines makes the non-covalent interactions more prevalent and allow for structural rearrangement. This stimulated the linear growth of worm-like aggregates. Full elimination of all five cysteines led to the transformation of worm-like aggregates into actual fibril structures (several hundreds of nanometres long) at pH 7.0. This understanding of the role of cysteine in protein-protein interactions will help to identify proteins and protein modifications to form functional aggregates at neutral pH.

Quantitative analysis of 3D food printing layer extrusion accuracy: Contextualizing automated image analysis with human evaluations: Quantifying 3D food printing accuracy.

3D food printing can customize food appearance, textures, and flavors to tailor to specific consumer needs. Current 3D food printing depends on trial-and-error optimization and experienced printer operators, which limits the adoption of the technology by general consumers. Digital image analysis can be applied to monitor the 3D printing process, quantify printing errors, and guide optimization of the printing process. We here propose an automated printing accuracy assessment tool based on layer-wise image analysis. Printing inaccuracies are quantified based on over- and under-extrusion with reference to the digital design. The measured defects are compared to human evaluations via an online survey to contextualize the errors and identify the most useful measurements to improve printing efficiency. The survey participants marked oozing and over-extrusion as inaccurate printing which matched the results obtained from automated image analysis. Although under-extrusion was also quantified by the more sensitive digital tool, the survey participants did not perceive consistent under-extrusion as inaccurate printing. The contextualized digital assessment tool provides useful estimations of printing accuracy and corrective actions to avoid printing defects. The digital monitoring approach may accelerate the consumer adoption of 3D food printing by improving the perceived accuracy and efficiency of customized food printing.

In silico modelling of protein digestion: A case study on solid/liquid and blended meals.

We present a dynamic, semi-mechanistic, compartmental protein digestion model to study the kinetics of protein digestion. The digestive system is described as a series of eight compartments: one for the stomach, one for the duodenum, two for the jejunum and four for the ileum. The digestive processes are described by a set of zero or first order differential equations. The model considers ingestion of a meal, secretion of gastric and pancreatic juices, protein hydrolysis, grinding, transit and amino acid absorption. The model was used to simulate protein digestion of a meal composed of a solid and a liquid phase or one where both phases are blended into a homogeneous phase. Luminal volumes and pH of gastric and duodenal contents were estimated for both meals. Further, gastric emptying is described as a function of the energy density of the bolus, instead of the more common mass action approach.

In vitro gastro-small intestinal digestion of conventional and mildly processed pea protein ingredients.

We report on the effect of processing, particularly heating, on the digestion dynamics of pea proteins using the standardised semi-dynamic in vitro digestion method. Fractions with native proteins were obtained by mild aqueous fractionation of pea flour. A commercial pea protein isolate was chosen as a benchmark. Heating dispersions of pea flour and mild protein fractions reduced the trypsin inhibitory activity to levels similar to that of the protein isolate. Protein-rich and non-soluble protein fractions were up to 18% better hydrolysed after being thermally denatured, particularly for proteins emptied later in the gastric phase. The degree of hydrolysis throughout the digestion was similar for these heated fractions and the conventional isolate. Further heating of the protein isolate reduced its digestibility as much as 9%. Protein solubility enhances the digestibility of native proteins, while heating aggregates the proteins, which ultimately reduces the achieved extent of hydrolysis from gastro-small intestinal enzymes.

Effect of Fractionation and Processing Conditions on the Digestibility of Plant Proteins as Food Ingredients.

Plant protein concentrates and isolates are used to produce alternatives to meat, dairy and eggs. Fractionation of ingredients and subsequent processing into food products modify the techno-functional and nutritional properties of proteins. The differences in composition and structure of plant proteins, in addition to the wide range of processing steps and conditions, can have ambivalent effects on protein digestibility. The objective of this review is to assess the current knowledge on the effect of processing of plant protein-rich ingredients on their digestibility. We obtained data on various fractionation conditions and processing after fractionation, including enzymatic hydrolysis, alkaline treatment, heating, high pressure, fermentation, complexation, extrusion, gelation, as well as oxidation and interactions with starch or fibre. We provide an overview of the effect of some processing steps for protein-rich ingredients from different crops, such as soybean, yellow pea, and lentil, among others. Some studies explored the effect of processing on the presence of antinutritional factors. A certain degree, and type, of processing can improve protein digestibility, while more extensive processing can be detrimental. We argue that processing, protein bioavailability and the digestibility of plant-based foods must be addressed in combination to truly improve the sustainability of the current food system.

Non-linear rheology reveals the importance of elasticity in meat and meat analogues.

The interest in plant-based meat analogues as an alternative to meat is currently growing. Rheological benchmarking is used to reveal how closely meat analogues resemble the original meat products. Texture maps and dissipation colour schemes were used to reveal similarities in and differences between rheological responses of meat and meat analogues (especially chicken analogues). Under heating, meat analogues differ in terms of their lower elasticity compared with heated meat. The changes caused by heating meat and meat analogues were different as well. Heating of meat resulted in a tougher and more elastic material, while heating has a minor effect on meat analogues. Future developments should therefore focus on routes to create more elasticity and possibly allow heating effects on texture to mimic meat characteristics even better.

Protein acidification and hydrolysis by pepsin ensure efficient trypsin-catalyzed hydrolysis.

Enzyme-catalysed hydrolysis is important in protein digestion. Protein hydrolysis is initiated by pepsin at low pH in the stomach. However, pepsin action and acidification happen simultaneously to gastric emptying, especially for liquid meals. Therefore, different extents of exposure to the gastric environment change the composition of the chyme that is emptied from the stomach into the small intestine over time. We assessed the susceptibility of a protein to trypsin-catalysed hydrolysis in the small intestine, depending on its pH and hydrolysis history, simulating chyme at different times after the onset of gastric emptying. Isothermal titration calorimetry was used to study the kinetics of pepsin and trypsin-catalysed hydrolysis. Bovine serum albumin (BSA) that was acidified and hydrolysed with pepsin, showed the highest extent and most efficient hydrolysis by trypsin. BSA in the chyme that would be first emptied from the stomach, virtually bypassing gastric acidity and peptic action, reduced trypsin-catalysed hydrolysis by up to 58% compared to the acidified, intact protein, and 77% less than the acidified, pepsin-hydrolysate. The least efficient substrate for trypsin-catalysed hydrolysis was the acidified, intact protein with a specificity constant (kcat/Km) nearly five times lower than that of the acidified, pepsin-hydrolysate. Our results illustrate the synergy between pepsin and trypsin hydrolysis, and indicate that gastric hydrolysis increases the efficiency of the subsequent trypsin-catalysed hydrolysis of a model protein in the small intestine.

Alternatives to Meat and Dairy.

The increasing size and affluence of the global population have led to a rising demand for high-protein foods such as dairy and meat. Because it will be impossible to supply sufficient protein to everyone solely with dairy and meat, we need to transition at least part of our diets toward protein foods that are more sustainable to produce. The best way to convince consumers to make this transition is to offer products that easily fit into their current habits and diets by mimicking the original foods. This review focuses on methods of creating an internal microstructure close to that of the animal-based originals. One can directly employ plant products, use intermediates such as cell factories, or grow cultured meat by using nutrients of plant origin. We discuss methods of creating high-quality alternatives to meat and dairy foods, describe their relative merits, and provide an outlook toward the future.

Separation of Fructose and Glucose via Nanofiltration in Presence of Fructooligosaccharides.

Fructose and glucose are commonly present together in mixtures and may need to be separated. Current separation methods for these isomers are complex and costly. Nanofiltration is a cost-effective method that has been widely used for separating carbohydrates of different sizes; however, it is not commonly used for such similar molecules. Here, we report the separation of fructose and glucose in a nanofiltration system in the presence of fructooligosaccharides (FOS). Experiments were performed using a pilot-scale filtration setup using a spiral wound nanofiltration membrane with molecular weight cutoff of 1 kDa. We observed three important factors that affected the separation: (1) separation of monosaccharides only occurred in the presence of FOS and became more effective when FOS dominated the solution; (2) better separation was achieved when the monosaccharides were mainly fructose; and (3) the presence of salt improved the separation only moderately. The rejection ratio (Rf/Rg) in a fructose/glucose mixture is 0.92. We reported a rejection ratio of 0.69, which was observed in a mixture of 50 g/L FOS with a fructose to glucose ratio of 4.43. The separation is hypothesized to occur due to selective transport in the FOS layer, resulting in a preferential binding towards fructose.

The effect of monovalent (Na+, K+) and divalent (Ca2+, Mg2+) cations on rapeseed oleosome (oil body) extraction and stability at pH 7.

Oleosomes are storage vehicles of TAGs in plant seeds. They are protected with a phospholipid-protein monolayer and extracted with alkaline aqueous media; however, pH adjustment intensifies the extraction process. Therefore, the aim of this work was to investigate the extraction mechanism of rapeseed oleosomes at pH 7 and at the presence of monovalent and divalent cations (Na+, K+, Mg2+, and Ca+2). The oleosome yield at pH 9.5 was 64 wt%, while the yield at pH 7 with H2O was just 43 wt.%. The presence of cations at pH 7, significantly enhanced the yield, with K+ giving the highest yield (64 wt.%). The cations affected the oleosome interface and their interactions. The presence of monovalent cations resulted in aggregation and minor coalescence, while divalent cations resulted in extensive coalescence. These results help to understand the interactions of oleosomes in their native matrix and design simple extraction processes at neutral conditions.

Analysis and modeling of enhanced green fluorescent protein diffusivity in whey protein gels.

During gastric digestion, hydrolysis of proteins by pepsin contributes largely to the breakdown of protein-rich food. We hypothesized that the effect of pepsin is limited by its diffusivity, which is co-determined by the food structure and the local pH in the food during digestion. To investigate the principle mechanism of enzyme diffusion in food matrices, we used enhanced green fluorescent protein (EGFP) as probe to study the diffusivity of proteins in whey protein isolate gels, using fluorescence correlation spectroscopy (FCS). Gels made with different ionic strength showed distinctive elastic moduli but did not show differences in diffusivity of EGFP. Some models for diffusion in hydrogels yield good description of the obtained data, and can approximate the enzyme diffusion in diverse food matrices. However, the enzyme pepsin is more complicated than the probe EGFP, to yield more accurate predictions, electrostatic and enzyme-substrate interaction also need to be considered.

Simultaneous Silicon Oxide Growth and Electrophoretic Deposition of Graphene Oxide.

During electrophoretic deposition of graphene oxide (GO) sheets on silicon substrates, not only deposition but also simultaneous anodic oxidation of the silicon substrate takes place, leading to a three-layered material. Scanning electron microscopy images reveal the presence of GO sheets on the silicon substrate, and this is also confirmed by X-ray photoelectron spectroscopy (XPS), albeit that the carbon portion increases with increasing emission angle, hinting at a thin carbon layer. With increasing applied potential and increasing conductivity of the GO solution, the carbon signal decreases, whereas the overall thickness of the added layer formed on top of the silicon substrate increases. Through XPS spectra in which the Si 2p peaks shifted under those conditions to 103-104 eV, we were able to conclude that significant amounts of oxygen are present, indicative of the formation of an oxide layer. This leads us to conclude that GO can be deposited using electrophoretic deposition, but that at the same time, silicon is oxidized, which may overshadow effects previously assigned to GO deposition.

Zwitterionic Polymer Modified Porous Carbon for High-Performance and Antifouling Capacitive Desalination.

Capacitive deionization (CDI) is an emerging technology for effective brackish water desalination to address fresh water scarcity. It is of great interest due to its high energy efficiency, environmental friendliness, and low-cost operation compared with traditional desalination technologies. However, electrode fouling, caused by dissolved organic matter and resulting in reduction of electrode electrosorption capacity and device lifespan, is an impediment to practical application of CDI. Herein, we report a novel salty water desalination electrode with excellent antifouling properties. The antifouling electrode is prepared by coating zwitterionic polymer brushes, i.e., poly(sulfobetaine methacrylate) (SBMA), on porous carbon (PC) via surface-initiated atom transfer radical polymerization. The successful coating of zwitterionic polymer on PC surface is confirmed by transmission electron microscopy, scanning electron microscopy, X-ray photoelectron spectroscopy, thermogravimetric analysis, and other characterizations. Coating with polySBMA did not affect the electrosorption capacity of PC electrodes and imparted antifouling properties (versus fouling by model foulant bovine serum albumin) during long-term salt removal tests (100 desalination/regeneration cycles). This is an important step toward practical application of capacitive deionization technology for brackish water desalination.

Interactions between acid and proteins under in vitro gastric condition - a theoretical and experimental quantification.

The gastric digestion of proteins is influenced by the pH and the gastric pH fluctuates after food consumption. However, the dynamics of gastric pH still need to be quantitatively understood. Proteins in food strongly influences the gastric pH. Therefore, we studied the interaction between acid and proteins, including the buffer reaction and the acid diffusion in protein gels. The buffer capacity of proteins stems from its content of ionizable amino acid side groups. Based on this, we set up a model and method to parameterize the buffer capacity of proteins. Moreover, the liberated carboxyl and amino groups during enzymatic hydrolysis of protein can also contribute to the buffer capacity. While we expected protons to diffuse faster than pepsin, we found that the penetration distance of acid is comparable to that of pepsin. The buffer reaction caused the acid to concentrate tenfold in the gel compared to the bulk acid concentration. Therefore, we postulated that the buffer reaction reduces acid diffusivity in gels.

Revisiting the enzymatic kinetics of pepsin using isothermal titration calorimetry.

Pepsin is the first protease that food proteins encounter in the digestive tract. However, most of the previous studies on the enzymatic kinetics of pepsin were based on the hydrolysis of small synthetic peptides, due to the limitations in methodology and the complexity of protein substrate. To better understand the role of pepsin in protein digestion, we used isothermal titration calorimetry to study the enzymatic kinetics of pepsin with bovine serum albumin as the substrate. We found that pepsin has a higher catalytic rate at lower pH, while its affinity to substrate is lower. At the same pH, pepsin has lower activity and affinity at higher ionic strengths. We found contrasting kinetic parameters for pepsin-catalyzed hydrolysis of bovine serum albumin and of small synthetic peptides. Time-dependent kinetics also showed that pepsin has lower efficiency towards intermediate peptides during hydrolysis.

The Effect of Gel Microstructure on Simulated Gastric Digestion of Protein Gels.

The objective of this study was to analyse the impact of the gel structure obtained by different heat-induced temperatures on the in vitro gastric digestibility at pH 2. To achieve this, gels were prepared from soy protein, pea protein, albumin from chicken egg white and whey protein isolate at varying temperatures (90, 120 and 140 °C) for 30 min. Gels were characterised prior to digestion via microstructure and SDS-PAGE analysis. Subsequently, the gastric digestion process was followed via the protein hydrolysis and HPSEC analysis up to 180 min. Peptides of different sizes (<5 kDa) were gradually formed during the digestion. Our results showed that gels induced at 140 °C were digested faster. The protein source and gelation temperature had great influence on the in vitro gastric protein digestibility.