Simulated gastro-intestinal digestion is widely employed in many fields of food and nutritional sciences, as conducting human trials are often costly, resource intensive, and ethically disputable. As a consequence, in vitro alternatives that determine endpoints such as the bioaccessibility of nutrients and non-nutrients or the digestibility of macronutrients (e.g. lipids, proteins and carbohydrates) are used for screening and building new hypotheses. Various digestion models have been proposed, often impeding the possibility to compare results across research teams. For example, a large variety of enzymes from different sources such as of porcine, rabbit or human origin have been used, differing in their activity and characterization. Differences in pH, mineral type, ionic strength and digestion time, which alter enzyme activity and other phenomena, may also considerably alter results. Other parameters such as the presence of phospholipids, individual enzymes such as gastric lipase and digestive emulsifiers vs. their mixtures (e.g. pancreatin and bile salts), and the ratio of food bolus to digestive fluids, have also been discussed at length. In the present consensus paper, within the COST Infogest network, we propose a general standardised and practical static digestion method based on physiologically relevant conditions that can be applied for various endpoints, which may be amended to accommodate further specific requirements. A frameset of parameters including the oral, gastric and small intestinal digestion are outlined and their relevance discussed in relation to available in vivo data and enzymes. This consensus paper will give a detailed protocol and a line-by-line, guidance, recommendations and justifications but also limitation of the proposed model. This harmonised static, in vitro digestion method for food should aid the production of more comparable data in the future.

https://hal.archives-ouvertes.fr/hal-01209876/document

A standardised static in vitro digestion method suitable for food – an international consensus

This paper describes the use of surface plasmon resonance spectroscopy and self-assembled monolayers (SAMs) of alkanethiols on gold to evaluate the ability of surfaces terminating in different combinations of charged groups to resist the nonspecific adsorption of proteins from aqueous buffer. Mixed SAMs formed from a 1:1 combination of a thiol terminated in a trimethylammonium group and a thiol terminated in a sulfonate group adsorbed less than 1% of a monolayer of two proteins with different characteristics:  fibrinogen and lysozyme. Single-component SAMs formed from thiols terminating in groups combining a positively charged moiety and a negatively charged moiety were also capable of resisting the adsorption of proteins. Single-component SAMs presenting single charges adsorbed nearly a full monolayer of protein. The amount of protein that adsorbed to mixed zwitterionic SAMs did not depend on the ionic strength or the pH of the buffer in which the protein was dissolved. The amount of protein that adsorbe...

Zwitterionic SAMs that Resist Nonspecific Adsorption of Protein from Aqueous Buffer

This contribution describes an experimentally straightforward procedure for preparing and screening surfaces for their ability to resist the adsorption of proteins from solution. For brevity, we call surfaces “protein resistant” when they are resistant to the adsorption of proteins from solution. We have used this procedure to identify several functional groups that had not previously been recognized as protein resistant. This work both identifies a number of functional groups that will be useful in designing resistance to the adsorption of biomolecules into devices used in sensing and in cell biology,1-3 and contributes to an understanding of the mechanism of action of protein resistant surfaces by correlating this property with molecular-scale structure.4-6 We combined self-assembled monolayers (SAMs)7,8 and surface plasmon resonance (SPR) spectroscopy9 into a system that enabled us to screen a number of functional groups rapidly for their ability to resist the adsorption of proteins. The surfaces were prepared by the “anhydride method” (Figure 1).10,11 This reaction generates a “mixed” SAM that comprises an ∼1:1 mixture of -CONRR′ and CO2H/CO2 groups.10,12 (We have not defined the state of the ionization of the CO2H groups in these SAMs.) The ease with which this class of mixed SAMs can be prepared by the anhydride method (relative to the synthesis of the functionalized alkanethiols HS(CH2)nR′ normally used for the preparation of singlecomponent SAMs) makes this route efficient for exploratory and screening work.11,13 We have examined the adsorption of two proteins to these surfaces: fibrinogen, a large (340 kD) blood plasma protein that adsorbs strongly to hydrophobic surfaces, and lysozyme, a small protein (14 kD, pI ) 12) that is positively charged under the conditions of our experiment (phosphate buffered saline, PBS, pH 7.4). Fibrinogen is used as a model for “sticky” serum proteins;11,14,15 lysozyme is often used in model studies of electrostatic adsorption of proteins to surfaces.16,17 Since lysozyme has a substantial net positive charge (Zp ) + 7.5 at pH 7.4, 100 mM KCl),18 it allowed us to examine attractive electrostatic interactions with CO2 groups on the surface. We have prepared more than 50 surfaces, each presenting a different functional group, using the anhydride procedure, and surveyed them for protein resistance.19 Table 1 summarizes selected results from this survey. The amount of protein adsorbed (∆RU ) change in response units) as measured by SPR was determined by subtracting the value of RU prior to the injection of protein from the value of RU measured 10 min after the completion of the protein injection; for clarity, these points are each labeled with a vertical dashed line in Figure 2. The value of ∆RU was used to calculate the percentage of a monolayer (%Monolayer) of that protein using eq 1.11,20 We define “%ML” according to eq 1 strictly to simplify the comparison between surfaces.

Surveying for Surfaces that Resist the Adsorption of Proteins

Additive manufacturing and digital fabrication bring new horizons to concrete and cement-based material construction. 3D printing inspired construction techniques that have recently been developed at laboratory scale for cement-based materials. This study aims to investigate the role of the structural build-up properties of cement-based materials in such a layer by layer construction technique. As construction progresses, the cement-based materials become harder with time. The mechanical strength of the cement-based materials must be sufficient to sustain the weight of the layers subsequently deposited. It follows that the comparison of the mechanical strength, which evolves with time (i.e. structural build-up), with the loading due to layers subsequently deposited, can be expected to provide the optimal rate of layer by layer construction. A theoretical framework has been developed to propose a method of optimization of the building rate, which is experimentally validated in a layer-wise built column.

https://link.springer.com/content/pdf/10.1617%2Fs11527-015-0571-0.pdf

Structural built-up of cement-based materials used for 3D-printing extrusion techniques

The archetypal feature of a viscoplastic fluid is its yield stress: If the material is not sufficiently stressed, it behaves like a solid, but once the yield stress is exceeded, the material flows like a fluid. Such behavior characterizes materials common in industries such as petroleum and chemical processing, cosmetics, and food processing and in geophysical fluid dynamics. The most common idealization of a viscoplastic fluid is the Bingham model, which has been widely used to rationalize experimental data, even though it is a crude oversimplification of true rheological behavior. The popularity of the model is in its apparent simplicity. Despite this, the sudden transition between solid-like behavior and flow introduces significant complications into the dynamics, which, as a result, has resisted much analysis. Over recent decades, theoretical developments, both analytical and computational, have provided a better understanding of the effect of the yield stress. Simultaneously, greater insight into the material behavior of real fluids has been afforded by advances in rheometry. These developments have primed us for a better understanding of the various applications in the natural and engineering sciences.

Yielding to Stress: Recent Developments in Viscoplastic Fluid Mechanics

The deformations and stresses during squeeze flows are evaluated for a wider class of materials than previously covered in articles on this subject. These include generalised Newtonian fluids, yield stress fluids, as well as elastic and viscoelastic materials. Wherever possible, results are given in a compact mathematical form. The effect of different boundary conditions (no slip, perfect slip and partial slip) and how these interact with different types of material behaviour to give a variety of macroscopic responses is also discussed. The significance of this in using squeeze flow as a rheometry method is highlighted and a state-of-the-art view of squeeze flow rheometry is given.

Squeeze flow theory and applications to rheometry: A review

Hydrogen-ion titrations were conducted for hen-egg-white lysozyme in solutions of potassium chloride over the range pH 2.5−11.5 and for ionic strengths to 2.0 M. The dependence of lysozyme's net proton charge, zp, on pH and ionic strength in potassium chloride solution is measured. From the ionic-strength dependence of zp, interactions of lysozyme with potassium and chloride ions are calculated using the molecular- thermodynamic theory of Fraaije and Lyklema.1 Lysozyme interacts preferentially with up to 12 chloride ions at pH 2.5. The observed dependence of ion−protein interactions on pH and ionic strength is explained in terms of electric-double-layer theory. New experimental pKa data are reported for 11 amino acids in potassium chloride solutions of ionic strength to 3.0 M.

/pdf/lysozyme-net-charge-and-ion-binding-in-concentrated-aqueous-3ye1ts1m0b.pdf

Lysozyme Net Charge and Ion Binding in Concentrated Aqueous Electrolyte Solutions

In the treatment of oropharyngeal dysphagia, the link between diagnosis and prescription of thickened liquids that are safe to swallow is not always straightforward. Frequently, the capacity to objectively assess and quantify the rheological properties of diagnostic test fluids and to select “rheologically equivalent” dietary products is missing. Perhaps sometimes the importance of an objective comparison is not fully appreciated because two liquids seem reasonably similar in a subjective comparison (e.g., flow from a spoon). The present study deals with some of these issues. Shear viscosity measurements were used to characterize the flow behavior of videofluoroscopic contrast agents and of thickened fluids prepared with commercial thickening agents. Effects of time and composition of the different fluids were analyzed regarding shear-rate-dependent viscosity. Nearly all materials tested showed a pronounced dependence of viscosity with shear rate (“shear thinning”). Results confirm that it is feasible (but not always straightforward) to “match” the viscosities of diagnostic fluids and thickened beverages if certain precautions are taken. For example, the time required to reach final viscosity levels can be significant for some thickeners, particularly when used with liquids containing contrast agents. It is recommend to use only diagnostic materials and thickening agents for which reliable viscosity data are available.

/pdf/matching-the-rheological-properties-of-videofluoroscopic-3fxy8uzh2e.pdf

Matching the Rheological Properties of Videofluoroscopic Contrast Agents and Thickened Liquid Prescriptions

Extrusion, structure and mechanical properties of complex starchy foams.

The understanding of how foods are digested and metabolised is essential to enable the design/selection of foods as part of a balanced diet. Essential to this endeavour is the development of appropriate biorelevant in vitro digestion tools. In this work, the influence of gastric pH profile on the in vitro digestion of mixtures of β-lactoglobulin (βlg) and xanthan gum prior to and after heat induced gelation was investigated. A conventional highly acidic (pH 1.9) gastric pH profile was compared to two dynamic gastric pH profiles (initial pH of 6.0 vs. 5.2 and H(+) secretion rates of 60 vs. 36 mmol h(-1)) designed to mimic the changes in gastric pH observed during clinical trials with high protein meals. In moving away from the pH 1.9 model, to a pH profile reflecting in vivo conditions, the initial rate and degree of protein digestion halved during the first 45 minutes. After 90 minutes of gastric digestion, all three pH profiles caused similar extents of protein digestion. Given that 50% gastric emptying times of (test) meals are in range of 30-90 min, it would seem highly relevant to use a dynamic pH gastric model rather than a pH 1.9 (USP) or pH 3 model (INFOGEST) in assessing the impact of food structuring approaches on protein digestion. The impact that heat induced gelation had on the degree of gel digestion by pepsin was also investigated. Surprisingly, it was found that heat induced gelation of βlg-xanthan mixtures at 70-90 °C for 20 minutes lead to a considerable decrease in the rate of proteolysis, which contrasts many studies of dispersed aggregates and gels of βlg alone whose heating accelerates pepsin activity due to unfolding. In the present case, the formation of a dense protein network created a fine pore structure which restricted pepsin access into the gel thereby slowing proteolysis. This work not only has implications for the in vitro assessment of protein digestion, but also highlights how protein digestion might be slowed, learnings that might have an influence on the design of foods as part of a satisfying balanced diet.

Jan Engmann

Papers

Squeeze flow theory and applications to rheometry: A review

Lysozyme Net Charge and Ion Binding in Concentrated Aqueous Electrolyte Solutions

Matching the Rheological Properties of Videofluoroscopic Contrast Agents and Thickened Liquid Prescriptions

Extrusion, structure and mechanical properties of complex starchy foams.

Impact of gastric pH profiles on the proteolytic digestion of mixed βlg-Xanthan biopolymer gels