- R.J.B.M. Delahaije (1)
- C. Forquenot de la Fortelle (1)
- H. Gruppen (1)
- S.L. Hefle (1)
- J.H. Ippel (1)
- H.H.J. Jongh de (1)
- M.M. Kool (1)
- S.J. Koppelman (1)
- S. Koutsopoulos (1)
- C.M.M. Lakemond (2)
- E. Linden van der (1)
- A.M. Matser (1)
- M.B.J. Meinders (1)
- C.P.M. Mierlo van (1)
- W. Norde (1)
- J. Oost van der (1)
- L.M.C. Sagis (1)
- G. Schiffart de (1)
- H.A. Schols (1)
- G. Sworn (1)
- A.M.M. Teeffelen Van (1)
- C. Veerman (1)
- C. Ven van der (1)
- P. Venema (1)
- R.W. Visschers (1)
- T. Vliet van (1)
- R.A.A. Vlooswijk (1)
- R.J. Vries de (1)
- M. Weijers (1)
- P.A. Wierenga (1)
Heat-Induced Aggregation of Whey Proteins in Aqueous Solutions below Their Isoelectric Point
Cornacchia, L. ; Forquenot de la Fortelle, C. ; Venema, P. - \ 2014
Journal of Agricultural and Food Chemistry 62 (2014). - ISSN 0021-8561 - p. 733 - 741.
bovine beta-lactoglobulin - disulfide bonds - interchange reactions - thermal-denaturation - alpha-lactalbumin - particulate gels - induced gelation - ionic-strength - ph 2 - solubility
Processing beverages containing high concentrations of globular proteins represents a technological challenge due to their instability during heating caused by protein aggregation and gelation. Aggregation of whey protein mixtures was investigated in aqueous model systems at pH 3.5, 4.0, and 4.5 at heating conditions resembling conventional industrial treatment (90 °C for 30 s). The extent of aggregation progressively decreased moving away from the pI. Protein aggregates became smaller and had a more open structure compared to higher pH values. Significant loss of protein dispersibility occurred at pH 4.0 and 4.5 above the denaturation T of whey protein (~70 °C), at which aggregation was caused by intermolecular hydrophobic interactions. Accessible thiol groups were detected in the heat-treated systems, with a higher intensity at higher pH and increasing extent of aggregation. Intermolecular -S-S- bonding played only a minor role in the aggregation at all conditions studied.
The influence of the primary and secondary xanthan structure on the enzymatic hydrolysis of the xanthan backbone
Kool, M.M. ; Schols, H.A. ; Delahaije, R.J.B.M. ; Sworn, G. ; Wierenga, P.A. ; Gruppen, H. - \ 2013
Carbohydrate Polymers 97 (2013)2. - ISSN 0144-8617 - p. 368 - 375.
bacterial polysaccharide xanthan - aqueous sodium-chloride - xanthomonas-campestris - extracellular polysaccharide - pyruvate substituents - thermal-denaturation - conformation change - behavior - acetyl - renaturation
Differently modified xanthans, varying in degree of acetylation and/or pyruvylation were incubated with the experimental cellulase mixture C1-G1 from Myceliophthora thermophila C1. The ionic strength and/or temperature of the xanthan solutions were varied, to obtain different xanthan conformations. The exact conformation at the selected incubation conditions was determined by circular dichroism. The xanthan degradation was analyzed by size exclusion chromatography. It was shown that at a fixed xanthan conformation, the backbone degradation by cellulases is equal for each type of xanthan. Complete backbone degradation is only obtained at a fully disordered conformation, indicating that only the secondary xanthan structure influences the final degree of hydrolysis by cellulases. It is thereby shown that, independently on the degree of substitution, xanthan can be completely hydrolyzed to oligosaccharides. These oligosaccharides can be used to further investigate the primary structure of different xanthans and to correlate the molecular structure to the xanthan functionalities.
Acid skim milk gels: The gelation process as affected by preheated pH
Lakemond, C.M.M. ; Vliet, T. van - \ 2008
International Dairy Journal 18 (2008)5. - ISSN 0958-6946 - p. 574 - 584.
glucono-delta-lactone - whey protein interactions - heat-induced interactions - casein micelles - rheological properties - beta-lactoglobulin - kappa-casein - thermal-denaturation - alpha-lactalbumin - dependent dissociation
The effect of preheating milk (10 min 80 [degree sign]C) at pH values from 6.20 to 6.90 on formation of acid skim milk gels was studied by dynamic oscillation measurements. Up to pH 6.65 a higher pH of heating (pHheating) resulted in a higher G'. Since below pH 4.9 the development of G'(pH)/G'(pH=4.9) and tan [delta] (pH) was similar we assume that the rearrangement processes happening below pH 4.9 were similar for all pHheating values studied. Based on data for tan [delta], G'(pH) and the pH of gel formation, it was hypothesized that the differences in moduli with pHheating stem from differences in the aggregation and rearrangement processes occurring before and just after gelation (pH>4.9). These differences are partly due to greater involvement of S-S interactions at higher pHheating. Furthermore, the soluble whey protein aggregates formed during preheating are in part not incorporated in the network, the extent depending on pH.
Kinetically controlled refolding of a heat denatured hyperthermostable protein
Koutsopoulos, S. ; Oost, J. van der; Norde, W. - \ 2007
FEBS Journal 274 (2007)22. - ISSN 1742-464X - p. 5915 - 5923.
archaeon pyrococcus-furiosus - crystal-structure - thermodynamic properties - thermal-denaturation - citrate synthase - stability - temperature - enzyme - water - dehydrogenase
The thermal denaturation of endo-ß-1,3-glucanase from the hyperthermophilic microorganism Pyrococcus furiosus was studied by calorimetry. The calorimetric profile revealed two transitions at 109 and 144¿°C, corresponding to protein denaturation and complete unfolding, respectively, as shown by circular dichroism and fluorescence spectroscopy data. Calorimetric studies also showed that the denatured state did not refold to the native state unless the cooling temperature rate was very slow. Furthermore, previously denatured protein samples gave well-resolved denaturation transition peaks and showed enzymatic activity after 3 and 9¿months of storage, indicating slow refolding to the native conformation over time.
High pressure versus heat treatments for pasteurisation and sterilisation of model emulsions
Ven, C. van der; Courvoisier, C. ; Matser, A.M. - \ 2007
Innovative Food Science and Emerging Technologies 8 (2007)2. - ISSN 1466-8564 - p. 230 - 236.
protein-stabilized emulsions - interchange reactions - emulsifying behavior - beta-lactoglobulin - whey proteins - sulfated polysaccharides - thermal-denaturation - induced gelation - water-interface - ionic-strength
Heat treatments can have considerable influence on the droplet size distribution of oil-in-water emulsions. In the present study, high-pressure (HP) pasteurisation and sterilisation were evaluated as alternatives for heat preservation of emulsions. HP conditions used were 600 MPa, 5 min, room temperature and 800 MPa, 5 min, 80 °C initial temperature, 115 °C maximum temperature for HP pasteurisation and HP sterilisation respectively. The effects on droplet size of these conditions were compared to heat treatments for whey protein isolate (WPI) and soy protein isolate (SPI) emulsions at two pH values and two ionic strengths. For WPI, also the effect of protein in the bulk phase was evaluated. Both HP and heat pasteurisation treatments resulted in similar or slightly decreased average droplet sizes compared to the untreated samples. For neutral SPI emulsions, heat sterilisation increased the average droplet size from 1.6 ¿m to 43.7 ¿m, while HP sterilisation resulted only in a small increase towards an average droplet size of 2.1 ¿m. The neutral WPI emulsions, except those with a high ionic strength, gave similar results with respect to the droplet size, showing that for neutral pH WPI or SPI emulsions HP sterilisation is preferable above heat sterilisation. Concerning the low pH WPI emulsions, the droplet sizes were unaffected after both heat and HP sterilisation
Identification of pitfalls in the analysis of heat capacity changes of beta-lactoglobulin A
Teeffelen, A.M.M. Van; Meinders, M.B.J. ; Jongh, H.H.J. de - \ 2005
International Journal of Biological Macromolecules 37 (2005)1-2. - ISSN 0141-8130 - p. 28 - 34.
monitor unfolding transitions - thermal-denaturation - fluorescence methods - protein stability - cold denaturation - thermodynamics - validity - curves - state - ph
Information on changes in heat capacity (¿Cp) of proteins upon unfolding is used frequently in literature to understand possible follow-up reactions of protein denaturation, like their aggregation propensity. This thermodynamic property is intrinsic to the protein's architecture and unfolding and should be independent of the approach used to evaluate it. However, for many proteins, the reported values for ¿Cp vary considerably. To identify whether the origin of these discrepancies lies within the experimental approach chosen and/or in the too simplified unfolding models used in the analysis of the data, we choose ß-lactoglobulin A, a relatively small protein, but disputed for its two-state unfolding, and established its ¿Cp from tryptophan fluorescence, near-UV circular dichroism and differential scanning calorimetric measurements. In view of the large variation for the obtained ¿Cp (between 3.2 and 10.1 ± 0.8 kJ/(mol K)), it is evident that: (1) the sensitivity of different approaches to the structural changes; (2) irreversibility of unfolding; (3) non-ideal two-state unfolding behaviour need to be considered prior to interpretation. While the first two points can be addressed by using multiple approaches, the applicability of the selected unfolding behaviour for the analysis is often less easy to establish. In this work, we illustrate that by checking the wavelength-dependence used to detect protein conformational changes a tool is provided that gives a direct insight in the validity of the interpretation in these studies. An experimentally validated determination of ¿Cp allows a more proper use for the mechanistic understanding of protein denaturation and its follow-up reactions, avoiding pitfalls in the interpretation.
Detection of soy proteins in processed foods: Literature overview and new experimental work
Koppelman, S.J. ; Lakemond, C.M.M. ; Vlooswijk, R.A.A. ; Hefle, S.L. - \ 2004
Journal of AOAC International 87 (2004)6. - ISSN 1060-3271 - p. 1398 - 1407.
linked immunosorbent-assay - heated meat-products - ionic-strength - monoclonal-antibodies - thermal-denaturation - soybean proteins - 11s globulin - glycine-max - functional-properties - gel-electrophoresis
Several tests for the detection of soy proteins in foods have been described in the literature, and some are commercially available. This article gives an overview of these methods and discusses the advantages and disadvantages of each individual method. Based on the conclusions of this inventory, an experimental approach was designed to improve the sensitivity of measuring soy protein in processed foods. The aimed sensitivity is 10 ppm (10 ¿g soy protein in 1 g solid sample), which is over 100-fold lower than presently available tests. The aimed sensitivity is this low because levels of food allergens at 10 ppm and above may provoke reactions in food allergic persons. Native soybean meal, soy protein isolate, soy protein concentrate, and textured soy flakes were used as test materials. Several extraction procedures were compared and a new method using high pH was selected. Polyclonal antibodies were raised in rabbits and goats, and immunopurified antibodies were used in sandwich and inhibition enzyme-linked immunosorbent assay (ELISA). Extraction at pH 12 resulted in good yields for all tested samples, both quantitatively (Bradford) and qualitatively by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Immunopurified rabbit antibodies against this extract used in a competition ELISA format resulted in a sensitive test with a detection limit of 0.02 ¿g/mL, corresponding to 0.4 ¿g/g (0.4 ppm) in food samples. Cross-reactivity with some main food ingredients was measured and appeared to be negative in all cases. The presently developed test is applicable for soy ingredients and soy-containing foods that are processed in different ways. The limit of quantitation is 1 ppm, which is an enormous improvement over earlier described methods.
Irreversible self-assembly of ovalbumin into fibrils and the resulting network rheology
Veerman, C. ; Schiffart, G. de; Sagis, L.M.C. ; Linden, E. van der - \ 2003
International Journal of Biological Macromolecules 33 (2003)1-3. - ISSN 0141-8130 - p. 121 - 127.
heat-induced networks - bovine serum-albumin - thermal-denaturation - aqueous-solutions - light-scattering - gels - ph - aggregation - proteins - microstructure
The self-assembly of ovalbumin into fibrils and resulting network properties were investigated at pH 2, as a function of ionic strength. Using transmission electron microscopy (TEM), the effect of ovalbumin concentration on the contour length was determined. The contour length was increasing with increasing ovalbumin concentration. TEM micrographs were made to investigate the effect of ionic strength on the contour length. In the measured ionic strength regime (0.01–0.035 M) fibrils of approximately equal length (±200 nm) were observed. TEM micrographs showed that the contour length of the fibrils, versus time after dilution, remained constant, which indicates that the self-assembly of ovalbumin is irreversible. Using the results of rheological measurements, we observed a decreasing critical percolation concentration with increasing ionic strength. We explain this result in terms of an adjusted random contact model for charged semiflexible fibrils. Hereby, this model has now been proven to be valid for fibril networks of ß-lg, BSA and, currently, for ovalbumin.
Multiple steps during the formation of beta-lactoglobulin fibrils
Arnaudov, L.N. ; Vries, R.J. de; Ippel, J.H. ; Mierlo, C.P.M. van - \ 2003
Biomacromolecules 4 (2003)6. - ISSN 1525-7797 - p. 1614 - 1622.
atomic-force microscopy - heat-induced gelation - particle gel formation - globular-proteins - induced denaturation - thermal-denaturation - amyloid fibrils - ovalbumin gels - ionic-strength - ph
In this study, the heat induced fibrilar aggregation of the whey protein beta-lactoglobulin is investigated at low pH and at low ionic strength. Under these circumstances, tapping mode atomic force microscopy results indicate that the fibrils formed have a periodic structure with a period of about 25 nm and a thickness of one or two protein monomers. Fibril formation is followed in situ using light scattering and proton NMR techniques. The dynamic light scattering results show that the fibrils that form after short heating periods (up to a few hours) disintegrate upon slow cooling, whereas fibrils that form during long heating periods do not disintegrate upon subsequent slow cooling. The NMR results show that even after prolonged heating an appreciable fraction of the protein molecules is incorporated into fibrils only when the beta-lactoglobulin concentration is above approximately 2.5 wt %. The data imply multiple steps during the heat induced formation of beta-lactoglobulin fibrils at low pH and at low ionic strength: (partly) denatured protein monomers are either incorporated into fibrils or form instead a low molecular weight complex that is incapable of forming fibrils. Fibril formation itself also involves (at least) two steps: the reversible formation of linear aggregates, followed by a slow process of "consolidation" after which the fibrils no longer disintegrate upon slow cooling.
Heat-induced denaturation and aggregation of ovalbumin at neutral pH described by irreversible first-order kinetics
Weijers, M. ; Barneveld, P.A. ; Cohen Stuart, M.A. ; Visschers, R.W. - \ 2003
Protein Science 12 (2003). - ISSN 0961-8368 - p. 2693 - 2703.
differential scanning calorimetry - protein denaturation - beta-lactoglobulin - thermal-denaturation - globular-proteins - light-scattering - disulfide bonds - dsc data - gelation - gels
The heat-induced denaturation kinetics of two different sources of ovalbumin at pH 7 was studied by chromatography and differential scanning calorimetry. The kinetics was found to be independent of protein concentration and salt concentration, but was strongly dependent on temperature. For highly pure ovalbumin, the decrease in nondenatured native protein showed first-order dependence. The activation energy obtained with different techniques varied between 430 and 490 kJ.mole(-1). First-order behavior was studied in detail using differential scanning calorimetry. The calorimetric traces were irreversible and highly scan rate-dependent. The shape of the thermograms as well as the scan rate dependence can be explained by assuming that the thermal denaturation takes place according to a simplified kinetic process N -->(k) D where N is the native state, D is denatured (or another final state) and k a first-order kinetic constant that changes with temperature, according to the Arrhenius equation. A kinetic model for the temperature-induced denaturation and aggregation of ovalbumin is presented. Commercially obtained ovalbumin was found to contain an intermediate- stable fraction (IS) of about 20% that was unable to form aggregates. The denaturation of this fraction did not satisfy first-order kinetics.