Design of whey protein nanostructures for incorporation and release of nutraceutical compounds in food

Whey proteins are widely used as nutritional and functional ingredients in formulated foods because they are relative inexpensive, generally recognized as safe (GRAS) ingredient and possess important biological, physical and chemical functionalities. Denaturation and aggregation behavior of these proteins is of particular relevance toward manufacture of novel nanostructures with a number of potential uses. When these processes are properly engineered and controlled, whey proteins may be formed into nanohydrogels, nanofibrils or nanotubes and be used as carrier of bioactive compounds. This review intends to discuss the latest understandings of nanoscale phenomena of whey protein denaturation and aggregation that may contribute for the design of protein nanostructures. Whey protein aggregation and gelation pathways under different processing and environmental conditions such as microwave heating, high voltage and moderate electrical fields, high pressure, temperature, pH and ionic strength were critically assessed. Moreover, several potential applications of nanohydrogels, nanofibrils and nanotubes for controlled release of nutraceutical compounds (e.g. probiotics, vitamins, antioxidants and peptides) were also included. Controlling the size of protein networks at nanoscale through application of different processing and environmental conditions can open perspectives for development of nanostructures with new or improved functionalities for incorporation and release of nutraceuticals in food matrices.Oscar L. Ramos, Ricardo N. Pereira and Clara Fuci~nos gratefully acknowledge their Post-Doctoral grants (SFRH/BPD/80766/2011, SFRH/BPD/ 81887/2011, and SFRH/BPD/87910/2012, respectively) to the Fundação para a Ciência e Tecnologia (FCT, Portugal). All authors thank the FCT Strategic Project PEst-OE/EQB/LA0023/2013 and the Project “BioEnv— Biotechnology and Bioengineering for a sustainable world”, REF. NORTE07-0124-FEDER-000048, co-funded by Programa Operacional Regional do Norte (ON.2–O Novo Norte), QREN, FEDER

In vitro impact of a whey protein isolate (WPI) and collagen hydrolysates (CHs) on B16F10 melanoma cells proliferation

Background: Porcine skin gelatine presented anti-tumoral effect on murine hepatoma cells (MH134), inducing programmed cell death (apoptosis). Whey proteins (mainly lactoferrin) have been investigated for cancer prevention and treatment. Objective: Investigation of the inhibitory capacity on melanoma cells (B16F10) proliferation and the influence on % distribution of cell cycle phases, in the presence of various concentrations of whey protein isolate (WPI), bovine Collagen hydrolysate (BCH) or its fractions. Methods: The permeate fraction BCH-P1 (molecular mass, MM 2.5 kDa) was further fractionated into five retentate fractions (R1-R5) by ultrafiltration membranes and into four fractions (F1-F4) by reverse phase chromatography. The permeate BCH-P1 and all its fractions were comparatively tested against a negative control (B16F10 cells + culture medium), and also against a positive control (B16F10 + culture medium + WPI). Results: The inhibitory concentrations for 50% of B16F10 cells (IC(50)) ranged from 0.19 to 156.9 mg/mL for all these proteins evaluated. The most inhibitory fractions of the BCH hydrolysate were BCH-P1 and F1-F4 with IC(50) concentrations below 1 mg/mL. Changes in cell cycle phases were characterized by a general decrease in the G2/M phase that emphasizes growth arrest, some increase in phase S (BCH-P1 and F4) but a strong increase in G0/G1 phase for BCH-P1 and F4. Caspase-3 expression increased significantly in all media containing F and R fractions, and also in the presence of BCH or WPI. Apoptosis was extremely high at low concentration (400 mu g/mL) of the F1-F3 fractions. Conclusion: It is suggested that a mechanism for tumorigenesis inhibition may involve the caspases cascade and apoptosis. (C) 2009 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.5615157Gelita South America, Cotia, S

Copper modulates the heat-induced sulfhydryl/disulfide interchange reactions of beta-Lactoglobulin

This study describes the effect of copper on the heat-denaturation/aggregation of beta-Lactoglobulin AB at neutral pH. The kinetics of disappearance of native beta-Lactoglobulin under different ionic strength and Cu(2+)/beta-Lactoglobulin molar ratio conditions were followed and the type of interactions (covalent or non-covalent) shared between non-native structures during the heating process were examined. On heating, the rate of disappearance of native beta-Lactoglobulin was accelerated by increasing the Cu(2+)/beta-Lactoglobulin molar ratio. Copper induces oxidation of the free sulfhydryl group of beta-Lactoglobulin resulting mainly in the formation of covalent dimers, which were further associated into large non-covalent aggregates under high ionic strength conditions. Characterisation of the beta-Lactoglobulin dimers reveals the existence of three different molecular species arising randomly (dimers A-A, A-B and B-B), in which tertiary structure was completely lost. The quantity of added copper constitutes a powerful way to control the heat-denaturation/aggregation process of beta-Lactoglobulin in particular regarding the relative proportion of covalent and non-covalent interactions into formed aggregates

Dry heating of whey proteins: Structures, Aggregation and Gelling properties

Dry heating is intensively used in pharmaceutical and food industry for viral and microbial decontamination of thermosensible proteins. It is rapidly developing as a pre-texturization process in food ingredient industry because it constitutes a means for improving the food proteins functionalities.[1] In this study, we investigated the chemical and structural changes that occur during dry heating under controlled physicochemical conditions and their consequences on the gelling properties of whey proteins. The water activity of the powder before dry heating and heat intensity (temperature/times) strongly accelerate the kinetic of denaturation/aggregation of whey proteins. The proteins composition and pH are the major factors that affect the structure of denatured and aggregated proteins. The kinetic of denaturation/aggregation of whey proteins was enhanced at pH 2.5 as compared to pH 6.5.[2] However, the presence of traces of lactose, as in case of commercially available WPI powders, modifies such kinetic order. At pH 2.5 the protein molecules were mainly linked together by intermolecular disulfide bonds, while at pH 6.5, covalent cross-links other than disulfide bonds were also involved in the formed aggregates. Stable non-native monomeric species of ?-Lg and ?-La exhibiting mass loss of 18 Da were generated during the course of dry heating, the proportion of which depends on pH. This mass loss has been attributed to N-term cyclisation for ?-La and to an internal cyclic imide formation for ?-Lg. Interestingly, the gelling properties were found to be highly dependent on the relative proportion of non-native monomers, soluble and insoluble aggregates that can be controlled by dry-heating pH. These results give new insight into the complex dry-heat induced modifications and aggregation pathways of whey proteins. They also give indications for a better control of physicochemical parameters of the powders at the industry to maintain the reproducibility of functionality of dry heated proteins ingredient

Dry heating of whey proteins: Structures, Aggregation and Gelling properties

Dry heating is intensively used in pharmaceutical and food industry for viral and microbial decontamination of thermosensible proteins. It is rapidly developing as a pre-texturization process in food ingredient industry because it constitutes a means for improving the food proteins functionalities.[1] In this study, we investigated the chemical and structural changes that occur during dry heating under controlled physicochemical conditions and their consequences on the gelling properties of whey proteins. The water activity of the powder before dry heating and heat intensity (temperature/times) strongly accelerate the kinetic of denaturation/aggregation of whey proteins. The proteins composition and pH are the major factors that affect the structure of denatured and aggregated proteins. The kinetic of denaturation/aggregation of whey proteins was enhanced at pH 2.5 as compared to pH 6.5.[2] However, the presence of traces of lactose, as in case of commercially available WPI powders, modifies such kinetic order. At pH 2.5 the protein molecules were mainly linked together by intermolecular disulfide bonds, while at pH 6.5, covalent cross-links other than disulfide bonds were also involved in the formed aggregates. Stable non-native monomeric species of ?-Lg and ?-La exhibiting mass loss of 18 Da were generated during the course of dry heating, the proportion of which depends on pH. This mass loss has been attributed to N-term cyclisation for ?-La and to an internal cyclic imide formation for ?-Lg. Interestingly, the gelling properties were found to be highly dependent on the relative proportion of non-native monomers, soluble and insoluble aggregates that can be controlled by dry-heating pH. These results give new insight into the complex dry-heat induced modifications and aggregation pathways of whey proteins. They also give indications for a better control of physicochemical parameters of the powders at the industry to maintain the reproducibility of functionality of dry heated proteins ingredient