Aggregation of whey protein hydrolysate using Alcalase 2.4 L.

Here, we describe peptide aggregation, which is also known as enzymatic protein resynthesis. Whey protein hydrolysate (WPH) is the starting material for assembling peptides. Analyses of the involved amino acids, intrinsic fluorescence, fluorescence phase diagram, secondary structure, turbidity, and surface hydrophobicity were performed to investigate the reaction process. The aggregation mechanism consists of two parts: 1) formation and 2) aggregation of the building blocks that form the ordered secondary β-sheet structure. Constructing the building blocks requires at least one intermediate state, which is formed after 0.5 hours. Non-synergistic changes in the secondary and tertiary structures then allow the intermediate state to emerge

Separation and Enrichment of Lectin from Zihua Snap-Bean (Phaseolus vulgaris) Seeds by PEG 600–Ammonium Sulfate Aqueous Two-Phase System

A fast and efficient method based on a polyethylene glycol (PEG) 600/(NH4)2SO4 aqueous two-phase system for extracting lectin from Zihua snap-bean (Phaseolus vulgaris) seeds was established. According to a Box–Behnken design (BBD), involving four factors at three levels each subjected to analysis of variance (ANOVA) and response surface analysis, the protein recovery and the purification factor of lectin in the top phase were used as the response values of the variance analysis to acquire the multivariate quadratic regression model. SDS–PAGE electrophoresis and the hemagglutination test were used to detect the distribution of lectin in the aqueous two-phase system (ATPS). The obtained data indicated that lectin was preferentially partitioned into the PEG-rich phase, and the ATPS, composed of 15% (NH4)2SO4 (w/w), 18% PEG 600 (w/w), 0.4 g/5 g NaCl and 1 mL crude extract, showed good selectivity for lectin when the pH value was 7.5. Under the optimal conditions, most of the lectin was assigned to the top phase in the ATPS, and the hemagglutination activity of the purified lectin in the top phase was 3.08 times that of the crude extract. Consequently, the PEG 600/(NH4)2SO4 aqueous two-phase system was an effective method for separating and enriching lectin directly from the crude extract of Zihua snap-bean seeds

Nitrogen-Doped Carbon Quantum Dots as Fluorescent Probes for Sensitive and Selective Detection of Nitrite

Nitrites are the upstream precursors of the carcinogenic nitrosamines, which are widely found in the natural environment and many food products. It is important to develop a simple and sensitive sensor for detecting nitrites. In this work, a fluorescence probe based on nitrogen-doped carbon quantum dots (N-CQDs) was developed for the sensitive and selective determination of nitrites. At pH 2, the fluorescence of N-CQDs can be selectively quenched by nitrite due to the fact N-nitroso compounds can be formed in the reaction of amide groups with nitrous acid, which results in fluorescence static quenching. Under optimal conditions, fluorescence intensity quenching upon addition of nitrite gives a satisfactory linear relationship covering the linear range of 0.2–20 μM, and the limit of detection (LOD) is 40 nM. Moreover, this method has been successfully applied to the determination of nitrites in tap water, which indicates its great potential for monitoring of nitrites in environmental samples

Separation of Antioxidant Peptides from Pepsin Hydrolysate of Whey Protein Isolate by ATPS of EOPO Co-polymer (UCON)/Phosphate

Abstract An aqueous two-phase system (ATPS) consisting of poly(ethylene glycol-ran-propylene glycol) monobutyl ether (UCON)/phosphate was developed for the separation of the antioxidant peptides from pepsin hydrolysate of Whey Protein Isolate (WPI). The efficiency of the separation was evaluated based on the DPPH radical scavenging activity, ABTS radical scavenging activity and ferric reducing antioxidant power (FRAP) of the separated peptides. The effects of some parameters on the partition of antioxidant peptides were investigated. An efficient separation of antioxidant peptides was achieved using ATPS with pH of 4.0, 4 mL of UCON solution (40%, w/w), 4 mL of KH2PO4 solution (15.5%, w/w), 2 mL of WPI hydrolysate and 0.40 g/10 mL of NaCl. Reversed-phase high-performance liquid chromatography (RP-HPLC), amino acid analyzer and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) were used to characterize the purified peptides separated by the ATPS. The peptides in top phase were less polar than those in bottom phase. More antioxidative and hydrophobic amino acids were extracted to the top phase of ATPS, and the peptides with the amino acid sequences with antioxidant activities moved to the top phase as well. In conclusion, antioxidant peptides were successfully separated from the WPI hydrolysate by UCON/phosphate ATPS

Targeting Delivery System for Lactobacillus Plantarum Based on Functionalized Electrospun Nanofibers

With the increased interest in information on gut microbes, people are realizing the benefits of probiotics to health, and new technologies to improve the viability of probiotics are still explored. However, most probiotics have poor resistance to adverse environments. In order to improve the viability of lactic acid bacteria, polylactic acid (PLA) nanofibers were prepared by coaxial electrospinning. The electrospinning voltage was 16 kV, and the distance between spinneret and collector was 15 cm. The feed rates of the shell and core solutions were 1.0 and 0.25 mL/h, respectively. The lactic acid bacteria were encapsulated in the coaxial electrospun nanofibers with PLA and fructooligosaccharides (FOS) as the shell materials. Scanning electron microscopy, transmission electron microscopy, and laser scanning confocal microscopy showed that lactic acid bacteria were encapsulated in the coaxial electrospun nanofibers successfully. The water contact angle test indicated that coaxial electrospun nanofiber films had good hydrophobicity. An in vitro simulated digestion test exhibited that the survival rate of lactic acid bacteria encapsulated in coaxial electrospun nanofiber films was more than 72%. This study proved that the viability of probiotics can be improved through encapsulation within coaxial electrospun PLA nanofibers and provided a novel approach for encapsulating bioactive substances

Separation and Enrichment of Antioxidant Peptides from Whey Protein Isolate Hydrolysate by Aqueous Two-Phase Extraction and Aqueous Two-Phase Flotation

At present, peptides are separated by molecular exclusion chromatography and liquid chromatography. A separation method is needed in any case, which can be scaled up for industrial scale. In this study, aqueous two-phase extraction (ATPE) and aqueous two-phase flotation (ATPF) were applied to separate and enrich antioxidant peptides from trypsin hydrolysates of whey protein isolates (WPI). The best experimental conditions were investigated, and the results were evaluated using the 2,2′-Azinobis-(3-ethylbenzthiazoline-6-sulphonate) (ABTS) free radical scavenging activity of the peptides-per-unit concentration and the recovery rate (Y) of peptides in the top phase of both ATPE and ATPF. Under optimal conditions, the Y and ABTS free radical scavenging activity per unit concentration in top phase of ATPE could reach 38.75% and 12.94%, respectively, and in ATPF could reach 11.71% and 29.18%, respectively. The purified peptides were characterized by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and reversed-phase high-performance liquid chromatography (RP-HPLC). PeptideCutter and PeptideMass were applied to analyze and calculate the peptide sequencing. KILDKVGINYWLAHK, VGINYWLAHKALCSEK, and TPEVDDEALEKFDKALK sequences having antioxidant activity were detected in the top phase of ATPE, and VGINYWLAHKALCSEK, KILLDKVGINYWLAHK, ILLDKVGINYWLAHK, IIAEKTKIPAVFK, KIIAEKTKIPAVFK, and VYVEELKPTPEGDLEILLQK sequences having antioxidant activity were detected in the top phase of ATPF. In conclusion, antioxidant peptides were successfully separated from the WPI hydrolysate by ATPE and ATPF; compared with ATPE, ATPF has superior specificity in separating antioxidant peptides

Eco-Innovation in Reusing Food By-Products: Separation of Ovalbumin from Salted Egg White Using Aqueous Two-Phase System of PEG 1000/(NH₄)₂SO₄

For the purpose of reducing pollution and the rational use of salted egg white, which is a byproduct of the manufacturing process of salted egg yolk, an aqueous two-phase system (ATPS) composed of polyethylene glycols (PEG 1000) and (NH4)2SO4 was investigated to selectively separate ovalbumin (OVA) from salted egg white. With the aim of optimizing the selective separation of OVA using ATPS, a response surface method (RSM) experiment was carried out on the basis of a single-factor experiment. The OVA was characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS⁻PAGE), reversed-phase high-performance liquid chromatography (RP-HPLC), liquid chromatography-nano electrospray ionization mass spectrometry (Nano LC-ESI-MS/MS), and Fourier transform infrared spectroscopy (FT-IR). Under the optimal conditions, the recovery yield of OVA through ATPS (Y) and the purity of OVA (P) could reach 89.25% and 96.28%, respectively. In conclusion, OVA was successfully separated from the salted egg white by PEG/(NH4)2SO4 ATPS

Curve-fitting results.

<p>Curve-fitting results.</p

Phase diagram of the aggregates according to incubation time.

<p>Phase diagram of the aggregates according to incubation time.</p