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

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.</p

Conformational changes of α-lactalbumin adsorbed at oil-water interfaces: interplay between protein structure and emulsion stability

The conformation and structural dimensions of α-lactalbumin (α-La) both in solution and adsorbed at oil-water interfaces of emulsions were investigated using synchrotron radiation circular dichroism (SRCD) spectroscopy, front-face tryptophan fluorescence (FFTF) spectroscopy, and dual polarization interferometry (DPI). The near-UV SRCD and the FFTF results demonstrated that the hydrophobic environment of the aromatic residues located in the hydrophobic core of native α-La was significantly altered upon adsorption, indicating the unfolding of the hydrophobic core of α-La upon adsorption. The far-UV SRCD results showed that adsorption of α-La at oil-water interfaces created a new non-native secondary structure that was more stable to thermally induced conformational changes. Specifically, the α-helical conformation increased from 29.9% in solution to 45.8% at the tricaprylin-water interface and to 58.5% at the hexadecane-water interface. However, the β-sheet structure decreased from 18.0% in solution to less than 10% at both oil-water interfaces. The DPI study showed that adsorption of α-La to a hydrophobic C18-water surface caused a change in the dimensions of α-La from the native globule-like shape (2.5-3.7 nm) to a compact/dense layer approximately 1.1 nm thick. Analysis of the colloidal stability of α-La stabilized emulsions showed that these emulsions were physically stable against droplet flocculation at elevated temperatures both in the absence and in the presence of 120 mM NaCl. In the absence of salt, the thermal stability of emulsions was due to the strong electrostatic repulsion provided by the adsorbed α-La layer, which was formed after the adsorption and structural rearrangement. In the presence of salt, although the electrostatic repulsion was reduced via electrostatic screening, heating did not induce strong and permanent droplet flocculation. The thermal stability of α-La stabilized emulsions in the presence of salt is a combined effect of the electrostatic repulsion and the lack of covalent disulfide interchange reactions. This study reports new information on the secondary and tertiary structural changes of α-La upon adsorption to oil-water interfaces. It also presents new results on the physical stability of α-La stabilized emulsions during heating and at moderate ionic strength (120 mM NaCl). The results broaden our understanding of the factors controlling protein structural change at emulsion interfaces and how this affects emulsion stability.Jiali Zhai, Søren V. Hoffmann, Li Day, Tzong-Hsien Lee, Mary Ann Augustin, Marie-Isabel Aguilar and Tim J. Wooste

Biodegradable biopolymer network structures to create delayed burst digestive release of encapsulated lipids

This study sought to investigate whether encapsulation of lipids in core-shell hydrogel structures of tailored shell porosity could create a delayed burst release of encapsulated lipids, and examined the underpinning mechanisms. We demonstrated that gastrointestinal digestion of core-shell structures resulted in a delay in the onset of lipid digestion, without affecting lipid digestion kinetics. Systematic increase in hydrogel protein content above 65 g/L lead to an exponential increase in digestive delay (250 min). Whilst an increase in xanthan content between 5 and 9 g/L lead to a modest decrease in digestive delay (40 min). Rheological investigations revealed a linear relationship between hydrogel storage modulus G′ and digestive breakdown delay (T1/2). Given that G’ is directly related to hydrogel mesh size, this result suggests that the main factor controlling the timing of digestive release is the average mesh size of the outer protein hydrogel. A kinetic model was created to describe the delayed burst release behaviour of encapsulated lipids and successfully predicted the influence of shell thickness, shell protein density on the timing of gastro-intestinal release (in vitro). By combining microstructural/rheological experiments with in vitro digestive studies we have understood the main factors controlling the digestive breakdown of hierarchical biopolymer hydrogels. We could successfully miniaturise these core-shell structures so they would easily empty from the stomach whilst maintaining programmable delayed burst release. We have created a novel family of core-shell hydrogel oral dosage forms for the delivery of poorly soluble drugs and the programmed delivery of lipids within the gut

Changes in beta-Lactoglobulin conformation at the oil/water interface of emulsions studied by synchrotron radiation circular dichroism spectroscopy

The structure of proteins at interfaces is a key factor determining the stability as well as organoleptic properties of food emulsions. While it is widely believed that proteins undergo conformational changes at interfaces, the measurement of these structural changes remains a significant challenge. In this study, the conformational changes of beta-lactoglobulin (beta-Lg) upon adsorption to the interface of hexadecane oil-in-water emulsions were investigated using synchrotron radiation circular dichroism (SRCD) spectroscopy. Far-UV SRCD spectra showed that adsorption of beta-Lg to the O/W interface caused a significant increase in non-native a-helix structure, accompanied by a concomitant loss of beta-sheet structure. Near-UV SRCD spectra revealed that a considerable disruption of beta-Lg tertiary structure occurred upon adsorption. Moreover, heat-induced changes to the non-native beta-Lg conformation at the oil/water interface were very small compared to the dramatic loss of beta-Lg secondary structure that occurred during heating in solution, suggesting that the interface has a stabilizing effect on the structure of non-native beta-Lg. Overall, our findings provide insight into the conformational behavior of proteins at oil/water interfaces and demonstrate the applicability of SRCD spectroscopy for measuring the conformation of adsorbed proteins in optically turbid emulsions

Soil respiration in a fire scar chronosequence of Canadian boreal jack pine forest

To fully understand the carbon (C) cycle impacts of forest fires, both C emissions during the fire and post-disturbance fluxes need to be considered. The latter are dominated by soil respiration (Rs), which is still subject to large uncertainties. This research investigates Rs in a boreal jack pine fire scar chronosequence at Sharpsand Creek, Ontario, Canada. During two field campaigns in 2006 and 2007, Rs was measured in a chronosequence of fire scars aged between 0 and 59 years since the last fire. Mean Rs per fire scar was adjusted for soil temperature (Ts) and soil moisture (Ms) (denoted RST,M). RST,M ranged from 0.56 μmol CO2/m2/s (32 years post fire) to 8.18 μmol CO2/m2/s (58 years post fire). The coefficient of variation (CV) of RST,M ranged from 20% (16 years post fire) to 56% (58 years post fire). Across the field site, there was a statistically highly significant exponential relationship between Rs adjusted for soil organic carbon (Cs) and Ts (P0.1) difference could be detected between recently burned (4 to 8 days post fire) and unburned young forest. There were significant differences in RST,M between recently burned (4 to 8 days post fire) scar age categories that differed in their burn history, with between-fire intervals of 32 vs. 16 years (P<0.001) and 32 vs 59 years (P=0.044). There was a highly significant exponential increase in RST,M with time since fire (r2=0.999; P=0.006) for the chronosequence 0, 16 and 59 years post fire, and for all these age categories, RST,M was significantly different from one another (P<0.05). The results of this study contribute to a better quantitative understanding of Rs in boreal jack pine fire scars and will facilitate improvements in C cycle modelling. Further work is needed in quantifying autotrophic and heterotrophic contributions to Rs in jack pine systems; in monitoring Rs for extended time periods after fire; and in measuring different fire-prone forest types

Soil surface CO[subscript 2] flux increases with successional time in a fire scar chronosequence of Canadian boreal jack pine forest

To fully understand the carbon (C) cycle impacts of forest fires, both C emissions during the fire and post-disturbance fluxes need to be considered. The latter are dominated by soil surface CO[subscript 2] flux (F[subscript s]), which is still subject to large uncertainties. Fire is generally regarded as the most important factor influencing succession in the boreal forest biome and fire dependant species such as jack pine are widespread. In May 2007, we took concurrent F[subscript s] and soil temperature (T[subscript s]) measurements in boreal jack pine fire scars aged between 0 and 59 years since fire. To allow comparisons between scars, we adjusted F[subscript s] for T[subscript s] (F[subscript s,superscript T]) using a Q[subscript 10] of 2. Mean F[subscript s,superscript T] ranged from 0.56 (± 0.30 sd) to 1.94 (± 0.74 sd) μmol CO[subscript 2] m[superscript −2] s[superscript −1]. Our results indicate a difference in mean F[subscript s,superscript T] between recently burned (4 to 8 days post fire) and non-burned mature (59 years since fire) forest (P < 0.001), though no difference was detected between recently burned (4 to 8 days post fire) and non-burned young (16 years since fire) forest (P = 0.785). There was a difference in mean F[subscript s,superscript T] between previously young (16 years since fire) and intermediate aged (32 years since fire) scars that were both subject to fire in 2007 (P < 0.001). However, there was no difference in mean F[subscript s,superscript T] between mature (59 years since fire) and intermediate aged (32 years since fire) scars that were both subjected to fire in 2007 (P = 0.226). Furthermore, there was no difference in mean F[subscript s,superscript T] between mature (59 years since fire) and young scars (16 years since fire) that were both subjected to fire in 2007 (P = 0.186). There was an increase in F[subscript s,superscript T] with time since fire for the chronosequence 0, 16 and 59 years post fire (P < 0.001). Our results lead us to hypothesise that the autotrophic:heterotrophic soil respiration ratio increases over post-fire successional time in boreal jack pine systems, though this should be explored in future research. The results of this study contribute to a better quantitative understanding of F[subscript s] in boreal jack pine fire scars and will facilitate meta-analyses of F[subscript s] in fire scar chronosequences