A New Method for Growth Factor Enrichment from Dairy Products by Electrodialysis with Filtration Membranes: The Major Impact of Raw Product Pretreatment.

This study is focused on fractionation of insulin-like growth factor I (IGF-I) and transforming growth factor-ß2 (TGF-ß2) using a new electro-based membrane process calledelectrodialysis with filtration membranes (EDFM). Before EDFM, different pretreatments were tested, and four pH conditions (4.25, 3.85, 3.45, and 3.05) were used during EDFM. It was demonstrated that a 1:1 dilution of defatted colostrum with deionized water to decrease mineral content followed by the preconcentration of GFs by UF is necessary and allow for these compounds to migrate to the recovery compartment during EDFM. MS analyses confirmed the migration, in low quantity, of only α-lactalbumin (α-la) and ß-lactoglobulin (ß-lg) from serocolostrum to the recovery compartment during EDFM. Consequently, the ratio of GFs to total protein in recovery compartment compared to that of feed serocolostrum solution was 60× higher at pH value 3.05, the optimal pH favoring the migration of IGF-I and TGF-ß2. Finally, these optimal conditions were tested on acid whey to also demonstrate the feasibility of the proposed process on one of the main by-products of the cheese industry; the ratio of GFs to total protein was 2.7× higher in recovery compartment than in feed acid whey solution, and only α-la migrated. The technology of GF enrichment for different dairy solutions by combining ultrafiltration and electrodialysis technologies was proposed for the first time.

The effect of heat treatment on the lactosylation of milk proteins.

Protein lactosylation is a significant modification that occurs during the heat treatment of dairy products, causing changes in proteins' physical-chemical and nutritional properties. Knowledge of the detailed lactosylation information on milk proteins under various heat treatments is important for selecting appropriate thermo-processing and identifying markers to monitor heat load in dairy products. In the present study, we used proteomics techniques to investigate lactosylated proteins under different heating temperatures. We observed a total of 123 lactosylated lysines in 65 proteins, with lactosylation even occurring in raw milk. The number of lactosylated lysines and proteins increased moderately at 75°C to 130°C, but dramatically at 140°C. We found that 6 out of 10, 9 out of 16, 6 out of 12, and 5 out of 15 lysine residues in κ-casein, ß-lactoglobulin, α-lactalbumin, and αS1-casein, respectively, were lactosylated under the applied heating treatment. Moreover, different lactosylation states of individual lysines and proteins can indicate the intensity of heating processes. Lactosylation of K14 in ß-lactoglobulin could distinguish pasteurized and UHT milk, while lactosylation of lactotransferrin can reflect moderate heat treatment of products.

Machine Learning Assisted Simultaneous Structural Profiling of Differently Charged Proteins in a Mycobacterium smegmatis Porin A (MspA) Electroosmotic Trap.

The nanopore is emerging as a means of single-molecule protein sensing. However, proteins demonstrate different charge properties, which complicates the design of a sensor that can achieve simultaneous sensing of differently charged proteins. In this work, we introduce an asymmetric electrolyte buffer combined with the Mycobacterium smegmatis porin A (MspA) nanopore to form an electroosmotic flow (EOF) trap. Apo- and holo-myoglobin, which differ in only a single heme, can be fully distinguished by this method. Direct discrimination of lysozyme, apo/holo-myoglobin, and the ACTR/NCBD protein complex, which are basic, neutral, and acidic proteins, respectively, was simultaneously achieved by the MspA EOF trap. To automate event classification, multiple event features were extracted to build a machine learning model, with which a 99.9% accuracy is achieved. The demonstrated method was also applied to identify single molecules of α-lactalbumin and ß-lactoglobulin directly from whey protein powder. This protein-sensing strategy is useful in direct recognition of a protein from a mixture, suggesting its prospective use in rapid and sensitive detection of biomarkers or real-time protein structural analysis.

Effects of a Novel High-Quality Protein Infant Formula on Energetic Efficiency and Tolerance: A Randomized Trial.

OBJECTIVES: Protein overfeeding in infants can have negative effects, such as diabetes and childhood obesity; key to reducing protein intake from formula is improving protein quality. The impact of a new infant formula [study formula (SF)] containing alpha-lactalbumin, lactoferrin, partially hydrolyzed whey, and whole milk on growth and tolerance compared to a commercial formula (CF) and a human milk reference arm was evaluated. METHODS: This randomized, double-blind trial included healthy, singleton, term infants, enrollment age ≤14 days. Primary outcome was mean daily weight gain. Secondary outcomes were anthropometrics, formula intake, serum amino acids, adverse events, gastrointestinal characteristics, and general disposition. RESULTS: Non-inferiority was demonstrated. There were no differences between the formula groups for z scores over time. Formula intake [-0.33 oz/kg/day, 95% confidence interval (CI): -0.66 to -0.01, P = 0.05] and mean protein intake (-0.13 g/kg/day, 95% CI: -0.26 to 0.00, P = 0.05) were lower in the SF infants, with higher serum essential amino acid concentrations (including tryptophan) compared to the CF infants. Energetic efficiency was 14.0% (95% CI: 8.3%, 19.7%), 13.0% (95% CI: 6.0%, 20.0%), and 18.1% (95% CI: 9.4%, 26.8%) higher for weight, length, and head circumference, respectively, in SF infants compared to the CF infants. SF infants had significantly fewer spit-ups and softer stool consistency than CF infants. CONCLUSIONS: The SF resulted in improved parent-reported gastrointestinal tolerance and more efficient growth with less daily formula and protein intake supporting that this novel formula may potentially reduce the metabolic burden of protein overfeeding associated with infant formula.

Identification of immunoglobulin E epitopes on major allergens from dairy products after digestion and transportation in vitro.

Dairy processing can alter the digestion stability and bioavailability of cow milk proteins in the gastrointestinal tract. However, analysis of stable linear epitopes on cow milk allergens that could enter into intestinal mucosal is limited. Thus, this study aimed to investigate the digestion and transportation properties and residual allergen epitopes entering into gastrointestinal mucosa of 3 commercial dairy products, including pasteurized milk (PM), ultra-heat-treated milk (UHTM), and dried skim milk (DSM). In this work, the digestive stability of the 3 kinds of dairy products has been performed in a standard multistep static digestion model in vitro and characterized by Tricine-SDS-polyacrylamide gel electrophoresis and reversed-phase HPLC. With respect to gastrointestinal digestion in vitro, the main allergens including ß-lactoglobulin (ß-LG), α-lactalbumin (α-LA), and caseins were degraded gradually, and the resistance peptides remained in the PM with a molecular weight of range from 3.4 to 5.0 kDa. Simultaneously, the potential allergenicity of the cow milk proteins was diminished gradually and is basically consistent after 60 min of gastrointestinal digestion. After gastrointestinal digestion, the remaining peptides were transported via an Ussing chamber and identified by liquid chromatography-MS/MS. By alignment, 10 epitopes peptides were identified from 16 stable peptides, including 5 peptides (AA 92-100, 125-135, 125-138, and 149-162) in ß-LG, 2 peptides in α-LA (AA 80-93 and 63-79), 2 peptides in αS1-casein (AA 84-90 and 125-132), and 1 peptide (AA 25-32) in αS2-casein were identified by dot-blotting mainly exist in UHTM and PM. This study demonstrates dairy processing can affect the digestion and transport characteristics of milk proteins and in turn alter epitope peptides release.

Analysis and comparison of key proteins in Maiwa yak and bovine milk using high-performance liquid chromatography mass spectrometry.

Yak milk is an essential and predominant food resource for Tibetan people for subsistence purposes and to combat altitude-induced challenges. Due to its unique qualities, yak milk has recently been gaining broader attention from consumers across China as well in other parts of the world. One of the key characteristics of yak milk is the protein content, which is about 40 to 60% higher than that of native bovine milk. In this work, a sensitive and reproducible high-throughput analytical method was developed employing both ultra high-performance liquid chromatography Orbitrap (Thermo Fisher Scientific) high-resolution accurate mass spectroscopy (UHPLC-HRAM-MS) and UHPLC coupled with triple quadrupole tandem MS (UHPLC-QqQ-MS) to simultaneously analyze 8 milk proteins. A total of 15 Maiwa yak milk samples and 15 bovine milk samples were qualitatively and quantitatively analyzed using targeted proteomics and compared for α-lactalbumin, ß-lactoglobulin, αS1-casein, αS2-casein, ß-casein, κ-casein, lactoferrin, and osteopontin. Peptides of ß-lactoglobulin were used to specifically distinguish yak and bovine milk. The results showed that this novel detection method could quantitatively detect these major and minor milk proteins with >0.99 linear correlation coefficient and a recovery rate between 90 and 120%, with relative standard deviations typically less than 10%. The data revealed that yak milk not only had higher overall milk protein content than bovine milk but higher lactoferrin and osteopontin contents as well. The lactoferrin content of yak milk was about 30% higher than that of bovine milk, and the osteopontin content of yak milk was nearly twice that of bovine milk. The application of this method demonstrates that UHPLC-HRAM-MS and UHPLC-QqQ-MS are suitable for high-throughput qualitative and quantitative analysis of major and minor proteins of yak and bovine milk.

Immunological Analysis of Isothiocyanate-Modified α-Lactalbumin Using High-Performance Thin Layer Chromatography.

Undirected modifications between food proteins and secondary plant metabolites can occur during food processing. The results of covalent interactions can alter the functional and biological properties of the proteins. The present work studied the extent of which covalent conjugation of the bioactive metabolite benzyl isothiocyanate (BITC; a glucosinolate breakdown product) to the whey protein α-lactalbumin affects the protein's allergenicity. Additional to the immunological analysis of native untreated and BITC-modified α-lactalbumin, the analysis of antigenic properties of proteolytically digested protein derivatives was also performed by high performance thin layer chromatography and immunostaining. As a result of the chemical modifications, structural changes in the protein molecule affected the allergenic properties. In this process, epitopes are destroyed or inactivated, but at the same time, buried epitopes can be exposed or newly formed, so that the net effect was an increase in allergenicity, in this case. Results from the tryptic hydrolysis suggest that BITC conjugation sterically hindered the cleavage sites for the enzyme, resulting in reduced digestibility and allergenicity. Residual antigenicity can be still present as short peptide fragments that provide epitopes. The desire to make food safer for allergy sufferers and to protect sensitized individuals from an allergenic reaction makes it clear that the detection of food antigens is mandatory; especially by considering protein interactions.

Direct separation and purification of α-lactalbumin from cow milk whey by aqueous two-phase flotation of thermo-sensitive polymer/phosphate.

BACKGROUND: α-lactalbumin (α-La) is of great interest to the industry as a result of its excellent functional properties and nutritional value. Aqueous two-phase flotation (ATPF) of thermo-sensitive polymer poly (ethylene glycol-ran-propylene glycol) monobutyl ether (UCON) and KH2 PO4 was applied to directly separate and purify α-La from milk whey, which was purposed to simplify the production process and reduced cost of production. RESULTS: The effect of ATPF composition and operating parameters on the flotation efficiency (E) and purity of α-La were investigated. The optimal conditions included 2 min of premixing time, 30 mL min-1 flow velocity and 20 min of flotation time, whereas the composition conditions comprised 35.0 mL 0.18 g mL-1 phosphate solution (containing 10% (cow milk whey/salt solution, v/v) cow milk whey, 50 ppm defoamer and 2 g NaCl) and 5.0 mL of 40% (w/w) UCON solution. Under the optimal conditions, E of α-La was 95.67 ± 1.04% and purity of α-La was 98.78 ± 1.19%. UCON was recovered by a thermally-induced phase separation and reused in next ATPF process without reducing E of α-La. Purified α-La was characterized by several key technologies. The results indicated that α-La in cow milk whey could be directly separated and purified by the ATPF and the purity was satisfactory. Moreover, it was suggested there was no obvious structure difference between the α-La separated by ATPF and the α-La standard. CONCLUSION: The present study enabled the recycling of UCON, providing an effective, economically viable and environmentally friendly approach for the separation and purification of protein. © 2021 Society of Chemical Industry.

Short communication: Caseins and α-lactalbumin content of camel milk (Camelus dromedarius) determined by capillary electrophoresis.

Camel milk has unique physical, nutritional, and technological properties when compared with other milks, especially bovine. Because proteins confer many of the properties of milk and its products, this study aimed to determine the proteins of camel milk, their correlations, and relative distribution. Raw milk samples were collected from 103 dromedary camels in the morning and evening. Capillary electrophoresis results showed wide variation in the concentrations (g/L) of proteins between samples as follows: α-lactalbumin, 0.3 to 2.9; αS1-casein, 2.4 to 10.3; αS2-casein, 0.3 to 3.9; ß-casein, 5.5 to 29.0; κ-casein, 0.1 to 2.4; unknown casein protein 1, 0.0 to 3.4; and unknown casein protein 2, 0.0 to 4.6. The range in percent composition of the 4 caseins were as follows: αS1, 12.7 to 35.3; αS2, 1.8 to 20.8; ß, 42.3 to 77.4; and κ, 0.6 to 17.4. The relative proportion of αS1-, αS2-, ß-, and κ-caseins in camel milk (26:4:67:3, wt/wt) differed from that of bovine milk (38:10:36:12, wt/wt). This difference might explain the dissimilarity between the 2 milks with respect to technical and nutritional properties.

Front-face fluorescence spectroscopy combined with chemometrics to detect high proteinaceous matter in milk and whey ultrafiltration permeate.

Proteinaceous matter can leak into the permeate stream during ultrafiltration (UF) of milk and whey and lead to financial losses. Although manufacturers can measure protein content in the finished permeate powders, there is currently no rapid monitoring tool during UF to identify protein leak. This study applied front-face fluorescence spectroscopy (FFFS) and chemometrics to identify the fluorophore of interest associated with the protein leak, develop predictive models to quantify true protein content, and classify the types of protein leak in permeate streams. Crude protein (CP), nonprotein nitrogen (NPN), true protein (TP), tryptone-equivalent peptide (TEP), α-lactalbumin (α-LA), and ß-lactoglobulin (ß-LG) contents were measured for 37 lots of whey permeate and 29 lots of milk permeate from commercial manufacturers. Whey permeate contained more TEP than did milk permeate, whereas milk permeate contained more α-LA and ß-LG than did whey permeate. The types of protein leak were thus identified for predictive model development. Based on excitation-emission matrix (EEM) of high- and low-TP permeates, tryptophan excitation spectra were collected for predictive model development, measuring TP content in permeate. With external validation, a useful model for quality control purposes was developed, with a root mean square error of prediction of 0.22% (dry basis) and a residual prediction deviation of 2.8. Moreover, classification models were developed using partial least square discriminant analysis. These classification methods can detect high TP level, high TEP level, and presence of α-LA or ß-LG with 83.3%, 84.8%, and 98.5% cross-validated accuracy, respectively. This method showed that FFFS and chemometrics can rapidly detect protein leaks and identify the types of protein leak in UF permeate. Implementation of this method in UF processing plants can reduce financial loss from protein leaks and maintain high-quality permeate production.

Electrochemical immunoassay for lactalbumin based on the use of ferrocene-modified gold nanoparticles and lysozyme-modified magnetic beads.

The authors describe a method for electrochemical determination of the breast cancer biomarker α-lactalbumin (α-LA) using disposable screen-printed carbon electrodes (SPCEs). Lysozyme-conjugated Fe3O4 nanoparticles (Lys-Fe3O4NPs) were used to capture α-LA on the surface of the SPCEs which then is trapped in an immunosandwich using secondary antibodies labeled with ferrocene-modified gold nanoparticles. The amperometric response of ferrocene (recorded at +0.1 V vs. silver pseudo-reference electrode) as well as the electrocatalytic activity of gold nanoparticles on the hydrogen evolution reaction (recorded at -1.0 V Vs Ag pseudo-reference electrode) was exploited to sense α-LA. A sensitive voltammetric response is observed, with (a) a sensitivity of 0.8789 µA·nM-1.cm-2, (b) a detection limit (LOD, at S/N = 3) as low as 0.07 ng·mL-1, and (c) linear response in the 0.75 to 630 ng mL-1 α-LA concentration range. The assay is selective and reproducible, and the SPCEs have good storage stability. The SPCEs were applied (a) to the analysis of (spiked) maternal milk, (b) of spiked serum from healthy and pregnant persons, and (c) of serum of patients suffering from breast cancer. Graphical abstract Schematic presentation of a sensitive electrochemical immunoassay platform based on ferrocene modified gold nanoparticles and lysozyme modified magnetic beads for the determination of alpha lactalbumin in human sera and breast milk by the amperometric response of ferrocene and hydrogen evolution reaction.

Analysis of Protein-Phenolic Compound Modifications Using Electrochemistry Coupled to Mass Spectrometry.

In the last decade, electrochemical oxidation coupled with mass spectrometry has been successfully used for the analysis of metabolic studies. The application focused in this study was to investigate the redox potential of different phenolic compounds such as the very prominent chlorogenic acid. Further, EC/ESI-MS was used as preparation technique for analyzing adduct formation between electrochemically oxidized phenolic compounds and food proteins, e.g., alpha-lactalbumin or peptides derived from a tryptic digestion. In the first step of this approach, two reactant solutions are combined and mixed: one contains the solution of the digested protein, and the other contains the phenolic compound of interest, which was, prior to the mixing process, electrochemically transformed to several oxidation products using a boron-doped diamond working electrode. As a result, a Michael-type addition led to covalent binding of the activated phenolic compounds to reactive protein/peptide side chains. In a follow-up approach, the reaction mix was further separated chromatographically and finally detected using ESI-HRMS. Compound-specific, electrochemical oxidation of phenolic acids was performed successfully, and various oxidation and reaction products with proteins/peptides were observed. Further optimization of the reaction (conditions) is required, as well as structural elucidation concerning the final adducts, which can be phenolic compound oligomers, but even more interestingly, quite complex mixtures of proteins and oxidation products.

Changes in the content of immunostimulating components of colostrum obtained from dairy cows at different levels of production.

BACKGROUND: Recent data indicate that almost 60% of colostrum samples do not have enough antibodies to provide adequate protection for the calf. Thus, the aim of this study was to determine the influence of the level of production of cows on the content of immunostimulating components of protein fraction of colostrum. The observations were conducted on two farms that keep dairy cows. One of the farms was selected to represent a typical performance level for the mass population of dairy cows in Poland, about 5500 kg of milk per cow per year (LI) and the second one was characterized by a higher level of production, about 8000 kg (HI). RESULTS: Based on the data obtained, 100% of the samples collected from LI had an immunoglobulin G (IgG) content >50 gL-1 , in the first collection. In the case of HI, 63% of the samples had an IgG content >50 gL-1 , with a distribution in the range of 10.7 to 72.7 gL-1 . The colostrum of LI cows was characterized by a higher content of lactoferrin, lysozyme, α-lactalbumin, and ß-lactoglobulin compared to the milk of HI cows. CONCLUSIONS: The immunostimulating components of colostrum declined rapidly in subsequent milkings or with the increased productivity of the cows. The concentration of IgG was negatively correlated with the performance level of cows, which means that high-producing cows had colostrum with a low concentration of IgG. © 2018 Society of Chemical Industry.

Competitive multicomponent anion exchange adsorption of proteins at the single molecule level.

We use single molecule spectroscopy to study a multicomponent, competitive protein adsorption system. Fluorescently-labeled α-lactalbumin proteins are super-resolved adsorbing to cationic anion-exchange ligands in the presence of a competitor, insulin. We find that the competitor reduces the number of binding events by blocking ligands throughout the observed measurement time while the single-site adsorption kinetics are unchanged.

Comparison of milk protein composition and rennet coagulation properties in native Swedish dairy cow breeds and high-yielding Swedish Red cows.

Recent studies have reported a very high frequency of noncoagulating milk in Swedish Red cows. The underlying factors are not fully understood. In this study, we explored rennet-induced coagulation properties and relative protein profiles in milk from native Swedish Mountain and Swedish Red Polled cows and compared them with a subset of noncoagulating (NC) and well-coagulating (WC) milk samples from modern Swedish Red cows. The native breeds displayed a very low prevalence of NC milk and superior milk coagulation properties compared with Swedish Red cows. The predominant variants in both native breeds were αS1-casein (αS1-CN) B, ß-CN A2 and ß-lactoglobulin (ß-LG) B. For κ-CN, the B variant was predominant in the Swedish Mountain cows, whereas the A variant was the most frequent in the Swedish Red Polled. The native breeds displayed similar protein composition, but varied in content of αS1-CN with 9 phosphorylated serines (9P) form. Within the Swedish Mountain cows, we observed a strong inverse correlation between the relative concentration of κ-CN and micelle size and a positive correlation between ionic calcium and gel firmness. For comparison, we investigated a subset of 29 NC and 28 WC milk samples, representing the extremes with regard to coagulation properties based on an initial screening of 395 Swedish Red cows. In Swedish Red, NC milk properties were found to be related to higher frequencies of ß-CN A2, κ-CN E and A variants, as well as ß-LG B, and the predominant composite genotype of ß- and κ-CN in the NC group was A2A2/AA. Generally, the A2A2/AA composite genotype was related to lower relative concentrations of κ-CN isoforms and higher relative concentrations of αS1-, αS2-, and ß-CN. Compared with the group of WC milk samples, NC milk contained a higher fraction of αS2-CN and α-lactalbumin (α-LA) but a lower fraction of αS1-CN 9P. In conclusion, milk from native Swedish breeds has good characteristics for cheese milk, which could be exploited in niche dairy products. In milk from Swedish Mountain cows, levels of ionic calcium seemed to be more important for rennet-induced gel firmness than variation in the relative protein profile. In Swedish Red, lower protein content as well as higher fraction of αS2-CN and lower fraction of αS1-CN 9P were related to NC milk. Further, a decrease in the frequency of the composite ß-κ-CN genotype A2A2/AA through selective breeding could have a positive effect on milk coagulation properties.

Development of a colloidal gold immunochromatographic strip assay for simple and fast detection of human α-lactalbumin in genetically modified cow milk.

The qualitative and quantitative declaration of food ingredients is important to consumers, especially for genetically modified food as it experiences a rapid increase in sales. In this study, we designed an accurate and rapid detection system using colloidal gold immunochromatographic strip assay (GICA) methods to detect genetically modified cow milk. First, we prepared 2 monoclonal antibodies for human α-lactalbumin (α-LA) and measured their antibody titers; the one with the higher titer was used for further experiments. Then, we found the optimal pH value and protein amount of GICA for detection of pure milk samples. The developed strips successfully detected genetically modified cow milk and non-modified cow milk. To determine the sensitivity of GICA, a quantitative ELISA system was used to determine the exact amount of α-LA, and then genetically modified milk was diluted at different rates to test the sensitivity of GICA; the sensitivity was 10 µg/mL. Our results demonstrated that the applied method was effective to detect human α-LA in cow milk.

Development of a real-time quantitative PCR assay using a TaqMan minor groove binder probe for the detection of α-lactalbumin in food.

Here, we report the development of a real-time PCR assay using a TaqMan minor groove binder (MGB, Genecore, NCBI: AF249896.1, 806-820) probe and primer sets designed to recognize the α-lactalbumin gene from the cow (Bos taurus). We evaluated the efficacy of this assay for detecting and quantifying cow α-lactalbumin in commercial foods. Our results demonstrated that the developed method was highly sensitive and showed high specificity for cow milk, with consistent detection of 0.05 ng of bovine DNA. We tested 42 commercial food samples with or without cow milk listed as an ingredient by using the developed assay. Among the 42 samples, 26 products that listed milk as an ingredient and 3 products might contain milk showed positive signals, whereas the other 9 products that did not contain milk and 4 products that might contain milk tested negative. Therefore, this method could be widely used for the detection of cow milk allergens in food.

Short communication: Selecting the most informative mid-infrared spectra wavenumbers to improve the accuracy of prediction models for detailed milk protein content.

The objective of this study was to investigate the ability of mid-infrared spectroscopy (MIRS) to predict protein fraction contents of bovine milk samples by applying uninformative variable elimination (UVE) procedure to select the most informative wavenumber variables before partial least squares (PLS) analysis. Reference values (n=114) of protein fractions were measured using reversed-phase HPLC and spectra were acquired through MilkoScan FT6000 (Foss Electric A/S, Hillerød, Denmark). Prediction models were built using the full data set and tested with a leave-one-out cross-validation. Compared with MIRS models developed using standard PLS, the UVE procedure reduced the number of wavenumber variables to be analyzed through PLS regression and improved the accuracy of prediction by 6.0 to 66.7%. Good predictions were obtained for total protein, total casein (CN), and α-CN, which included αS1- and αS2-CN; moderately accurate predictions were observed for κ-CN and total whey protein; and unsatisfactory results were obtained for ß-CN, α-lactalbumin, and ß-lactoglobulin. Results indicated that UVE combined with PLS is a valid approach to enhance the accuracy of MIRS prediction models for milk protein fractions.

Effect of processing on polyamine content and bioactive peptides released after in vitro gastrointestinal digestion of infant formulas.

This study examined the influence of processing on polyamines and peptide release after the digestion of a commercial infant formula designed for children during the first months of life. Polyamine oxidase activity was not suppressed during the manufacturing process, which implicates that polyamine concentrations were reduced over time and during infant formula self-life. In gel electrophoresis, in vitro gastrointestinal digestion of samples with reduced amount of enzymes and time of digestion shows an increase in protein digestibility, reflected in the increase in nonprotein nitrogen after digestion and the disappearance of ß-lactoglobulin and α-lactalbumin bands in gel electrophoresis. Depending on the sample, between 22 and 87 peptides were identified after gastrointestinal digestion. A peptide from ß-casein f(98-105) with the sequence VKEAMAPK and antioxidant activity appeared in all of the samples. Other peptides with antioxidant, immunomodulatory, and antimicrobial activities were frequently found, which could have an effect on infant health. The present study confirms that the infant formula manufacturing process determines the polyamine content and peptidic profile after digestion of the infant formula. Because compositional dissimilarity between human milk and infant formula in polyamines and proteins could be responsible for some of the differences in health reported between breast-fed and formula-fed children, these changes must be taken into consideration because they may have a great effect on infant nutrition and development.

Distyrylbenzene-aldehydes: identification of proteins in water.

Three different, water soluble, aldehyde-appended distyrylbenzene (DSB) derivatives were prepared. Their interaction with different albumin variants (human, porcine, bovine, lactalbumin, ovalbumin) was investigated (pH 11). All three fluorophores exhibit graded, protein-dependent fluorescence turn-on at slightly differing wavelengths. Linear discriminant analysis (LDA) differentiated all of the investigated albumins and was used to discern commercially available protein shakes. The three DSB derivatives barely react with the constituting amino acids but cysteine. In the proteins significant fluorescence signals are generated, probably due to a combination of imine/N,S-aminal formation and hydrophobic interactions between the DSBs and the proteins.