Influence of pulsed light treatment on the aggregation of whey protein isolate.

The effect of pulsed light (PL) on the aggregation of whey protein isolate (WPI) solutions was investigated. PL fluence values from 4 to 16J/cm2 were used to treat WPI (1% w/v) solutions in sodium phosphate buffer (pH=7.5). Whey protein structural modification and aggregation were assessed through the determination of free SH-groups and UV-absorption spectra. Additionally, covalent and non-covalently linked protein-protein interactions were identified through the measurement of turbidity, aggregation index, particle size distribution, and SDS-PAGE. WPI upon PL treatment showed structural changes as demonstrated by the immediate increase of free SH-group content (unfolding) and the subsequent formation of a small fraction of aggregation of unfolded proteins, due to both hydrophobic interactions and the formation of disulphide bonds. Turbidity, mean particle size, and aggregation index increased in samples treated at PL fluence from 4 to 16J/cm2. Furthermore, particle size distribution analysis of samples treated at higher fluence indicated that WPI dimer dissociation and formation of larger particles were likely to occur. The association of intermediate and larger protein molecules as well as the formation of soluble aggregates between ß-lactoglobulin and α-lactalbumin were also observed in gel electrophoresis analysis. In conclusion, the results of this investigation demonstrated the potential of PL treatments to induce protein denaturation, with a minimal formation of soluble protein aggregates.

Development of iron-rich whey protein hydrogels following application of ohmic heating - Effects of moderate electric fields.

The influence that ohmic heating technology and its associated moderate electric fields (MEF) have upon production of whey protein isolate cold-set gels mediated by iron addition was investigated. Results have shown that combining heating treatments (90°C, 5min) with different MEF intensities let hydrogels with distinctive micro and macro properties - i.e. particle size distribution, physical stability, rheological behavior and microstructure. Resulting hydrogels were characterized (at nano-scale) by an intensity-weighted mean particle diameter of 145nm, a volume mean of 240nm. Optimal conditions for production of stable whey protein gels were attained when ohmic heating treatment at a MEF of 3V∙cm-1 was combined with a cold gelation step using 33mmol∙L-1 of Fe2+. The consistency index of hydrogels correlated negatively to MEF intensity, but a shear thickening behavior was observed when MEF intensity was increased up to 10V∙cm-1. According to transmission electron microscopy, ohmic heating gave rise to a more homogenous and compact fine-stranded whey protein-iron microstructure. Ohmic heating appears to be a promising technique, suitable to tailor properties of whey protein gels and with potential for development of innovative functional foods.

Effect of dynamic high pressure on functional and structural properties of bovine serum albumin.

Dynamic high pressure (DHP) has been investigated as an innovative suitable method to induce protein modifications. This work evaluated the effect of DHP (up to three passes at 100, 150 and 200MPa, with an inlet temperature of 20°C) on functional and structural properties of bovine serum albumin (BSA). Results indicated that DHP process applied up to an energy limit of 100MPa increased the protein foaming capacity (FC) (p<0.05 - increase up to 63% after 1 pass at 100MPa) and the utilization of multiple passes at high pressure promoted a reduction in this property (p<0.05 - reduction up to 31.6% after 3 passes at 200MPa). Similar results were observed for sulfhydryl group, indicating an influence of free thiol groups on FC. Complementarily, DHP process promoted an increase of proteins particles size, suggesting a new rearrangement of their conformational structure. DHP did not affect tryptophan microenvironment in BSA; however, this process induced the rearrangement of secondary structure elements. In the first cycle, the pressure increase resulted in a loss of secondary structure, while in the second and third cycles the DHP process resulted in the gain of secondary structure elements. These results indicated that the second and third passes triggered a molecular rearrangement of the protein structure, giving rise to a novel and more stable conformational state. This conclusion was also supported by thermal unfolding studies (melting temperature reduction from 67.5 to 54.6°C after 1 pass at 200MPa), in which the additional cycles of DHP caused the occurrence of an initial denaturation at high temperatures, compared to the first cycle.

Effect of high hydrostatic pressure on the enzymatic hydrolysis of bovine serum albumin.

BACKGROUND: The extent of enzymatic proteolysis mainly depends on accessibility of the peptide bonds, which stabilize the protein structure. The high hydrostatic pressure (HHP) process is able to induce, at certain operating conditions, protein displacement, thus suggesting that this technology can be used to modify protein resistance to the enzymatic attack. This work aims at investigating the mechanism of enzymatic hydrolysis assisted by HHP performed under different processing conditions (pressure level, treatment time). Bovine serum albumin was selected for the experiments, solubilized in sodium phosphate buffer (25 mg mL-1 , pH 7.5) with α-chymotrypsin or trypsin (E/S ratio = 1/10) and HPP treatment (100-500 MPa, 15-25 min). RESULTS: HHP treatment enhanced the extent of the hydrolysis reaction of globular proteins, being more effective than conventional hydrolysis. At HHP treatment conditions maximizing the protein unfolding, the hydrolysis degree of proteins was increased as a consequence of the increased exposure of peptide bonds to the attack of proteolytic enzymes. The maximum hydrolysis degree (10% and 7% respectively for the samples hydrolyzed with α-chymotrypsin and trypsin) was observed for the samples processed at 400 MPa for 25 min. At pressure levels higher than 400 MPa the formation of aggregates was likely to occur; thus the degree of hydrolysis decreased. CONCLUSION: Protein unfolding represents the key factor controlling the efficiency of HHP-assisted hydrolysis treatments. The peptide produced under high pressure showed lower dimensions and a different structure with respect to those of the hydrolysates obtained when the hydrolysis was carried out at atmospheric pressure, thus opening new frontiers of application in food science and nutrition. © 2016 Society of Chemical Industry.

Effect of pulsed light treatment on structural and functional properties of whey protein isolate.

This work aimed at investigating the effects of Pulsed Light (PL) processing at different fluences (from 4 to 16J/cm2) on the structure and functional properties of Whey Protein Isolate (WPI) solution. The determination of the free and total sulfhydryl (SH) groups was used to detect the variation of WPI tertiary and quaternary structure. Additionally, PL-induced changes in secondary structure were determined by FT-IR spectroscopy and the differential scanning calorimetry (DSC), and primary structure by carbonyl content. The experimental data demonstrated that PL treatments increased the concentration of total and free sulfhydryl groups and protein carbonyls. A decrease of the denaturation temperature and enthalpy ratio with increasing the intensity of PL treatments was observed in DSC measurements. Small but significant changes in the secondary structure of PL treated WPI solution were also taking place and detected. The extent of whey protein structure modifications was fluence dependent. The results of this investigation demonstrated the potential of PL treatments to induce dissociation and partial unfolding of WPI, thus improving some of their functional properties, such as solubility and foaming ability.

Pasteurization of fruit juices by means of a pulsed high pressure process.

The use of pulsed high hydrostatic pressure was investigated as a possible approach to stabilize foodstuffs. The objective of this article was to investigate the effect of the main processing variables (pressure [150 to 300 MPa], temperature levels [25 to 50 degrees C], and pulse number [1 to 10]) on the sanitation of nonpasteurized clear Annurca apple juice as well as freshly-squeezed clear orange juice. The aim of the article was the optimization of the process parameters in step-wise pressure treatment (pressure holding time of each pulse: 60 s, compression rate: 10.5 MPa/s, decompression time: 2 to 5s). The shelf life of the samples, processed at optimized conditions, was evaluated in terms of microbiological stability and quality retention. According to our experimental results, the efficiency of pulsed high pressure processes depends on the combination of pulse holding time and number of pulses. The pulsed high pressure cycles have no additive or synergetic effect on microbial count. The efficacy of the single pulses decreases with the increase of the pulse number and pressure level. Therefore the first pulse cycle is more effective than the following ones. By coupling moderate heating to high pressure, the lethality of the process increases but thermal degradation of the products can be detected. The optimization of the process condition thus results in a compromise between the reduction of the pressure value, due to the synergetic temperature action, and the achievement of quality of the final production. The juices processed under optimal processing conditions show a minimum shelf life of 21 d at a storage temperature of 4 degrees C.