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At present in the Russian Federation fortified food products have gained extensive promotion as the newest and promising direction in the food industry. Almost every dairy processing enterprise has a line of fortified products in its product range. Vitamins, antioxidants, vitamin-like substances, flavoring components, micro and macro elements, phospholipids, prebiotics, probiotics, essential amino acids, polyunsaturated fatty acids can be used as enriching ingredients. On the other hand, the problem of targeted and economically profitable use of secondary milk processing resources is a significant problem. Production of cottage cheese, cheese and casein produces a liquid by-product, which is one of the largest additional sources of dietary protein and lactose as an important source of carbohydrates. The aim of the research is to develop whey fermented milk drink. The objectives of the research are to determine the effect of Jerusalem artichoke syrup on the organoleptic characteristics of the developed fermented milk drink; to optimize empirically the recipe composition of the developed fermented milk drink; to select experimentally the species composition of the starter micro flora of the developed fermented milk drink. The objects of the research are milk serum, Jerusalem artichoke syrup, ground ginger and cinnamon, starter culture on pure cultures of lactic acid microorganisms consisting of Str. thermophilus strains, viscous strains and Lbm. Bulgaricus and B. bifidum 791 in a ratio of 2:1:1. It can be concluded that the use of flavored fillers in the technology of whey fermented milk drink makes it possible to obtain a new product with competitive characteristics.

The objective of this research is to determine whether intramammary antibiotics with complementary acupuncture can reduce bovine mammary inflammation due to subclinical mastitis. Lactating cows were selected based on milk with a somatic cell count (SCC) greater than 500,000 cells/ml. Pre- and post-treatment milk samples were collected to determine SCC, aerobic bacterial content, milk ion conductivity, total protein, lactate dehydrogenase (LDH) and N-acetyl-beta-D-glucosaminidase (NAGase) concentrations. Milk serum was prepared from milk samples by double centrifugation. Concentrations of LDH and NAGase were determined using commercial enzyme-linked immunosorbent assays. Cows being treated with intramammary antibiotics were separated by random assignment to the acupuncture group (n = 10) and a no-acupuncture (control) group (n = 9). Both the acupuncture and control group were restrained for 30 min in a head catch 12 hr apart for a total of four times. For front quarters affected by subclinical mastitis, the acupuncture points used were spleen (SP) 12, SP 17, SP 18, SP 21, stomach (ST) 18 and conception vessel (CV) 12. For rear quarters affected by subclinical mastitis, the acupuncture points used were bladder (BL) 30, BL 30-1, BL 49, kidney (KI) 10, conception vessel (CV) 2 and CV 3. All parameters were compared using a Student t test. Significance was defined as p < .05. Compared to control cows, complementary acupuncture treatment reduced NAGase enzymatic activity in quarters of cows with subclinical mastitis. The reduction in NAGase suggests that complementary acupuncture treatment may be associated with healing of the damaged mammary epithelial cells, which are the primary source of NAGase activity in milk serum.

During whey powder production, the feed is subjected to several heat treatments which can cause lactosylation of proteins. In this study, lactosylation of whey proteins was evaluated in spray-dried powders before and after storage by varying the native protein fraction as well as the serum protein/lactose ratio in the powders. The lactosylation of native α-lactalbumin and β-lactoglobulin in the powders before storage was not affected to a large extent by the protein denaturation or if the feed had been heat treated in a high or low lactose environment. After storage (relative humidity of 23.5%, 30 °C, 25 days), the kinetic of lactosylation tended to increase with increasing native protein fraction and bulk protein content in the powders. An explanation could be that proteins dissolved in the lactose glassy structure might have a lower reactivity, while proteins present in the protein glassy structure with dissolved lactose may display higher lactosylation reactivity. (Less)

In this study, milk protein isolate (MPI) and micellar casein concentrate (MCC) were obtained with ultrafiltration (UF) and microfiltration (MF), respectively. The objective of this work was to evaluate the effect of diafiltration (DF), the addition of water during filtration, on the colloidal dissociation of casein micelles. Two levels of DF were applied after an initial concentration step to 2x the volume fraction, at a total protein concentration of about 7% (w/v). Under these conditions, while permeation of protein was still measured in the permeates, calcium ion transmission reached a plateau. The molecular details of the serum phases were studied throughout the filtration process. A comparison of serum phases with and without whey protein (UF vs MF) was carried out by examining the two processes in parallel, applying similar filtration conditions. Final 4x concentrates (based on initial volume of milk) were also compared. Viscosity was higher for UF than the corresponding MF concentrates, is spite of the higher level of colloidal protein present in the MF concentrates, which would suggest a change in the hydration or the volume/weight ratio (voluminosity) of casein micelles in MCC retentates. Centrifugal supernatants contained casein proteins both in MPC and MCC retentates. There were no statistically significant changes in the apparent hydrodynamic diameter of the casein micelles when measured by dynamic light scattering, in the original milk serum. A two dimensional chromatography was carried out on the soluble aggregates to determine differences in the amount of individual caseins. The aggregates showed only traces of whey proteins. The composition of the final concentrates, albeit similar in total proteins, showed profound differences in ionic strength and soluble proteins, suggesting that these parameters may be used as an indication of ingredient quality and functionality.

Orientador: Cesar Costapinto Santana Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia Quimica Resumo: O crescente interesse e aplicações dos produtos biotecnológicos vem aumentando o desenvolvimento de novos processos de recuperação e purificação de proteínas. O soro de leite bovino, obtido da manufatura da caseína para a produção de queijo, é em sua maioria descartado em mananciais de água, causando sérios problemas ambientais devido a sua alta demanda biológica de oxigênio (DBO). As proteínas presentes no lactosoro apresentam um excelente valor nutritivo e farmacológico, porém, o seu uso no enriquecimento de produtos alimentícios é limitado devido a baixa concentração destas proteínas. As principais proteínas do lactosoro são: ~-Iactoglobulinas, a-Iactalbumina, albuminas de soro bovino, imunoglobulinas, lactoperoxidase, lactoferrina, lisozima e outras proteínas de menor proporção, que apresentam um alto valor agregado. A a-Iactalbumina atua no organismo estimulando os agentes do sistema imunológico por proporcionar a elevação de glutationa em vários órgãos e no sangue, resultando em benefícios para pacientes portadores de doenças degenerativas como os males de Parkinson e Alzheimer, câncer e AIDS. Neste trabalho foi estudado o processo de separação da a-Iactalbumina, através de técnicas cromatográficas empregando a metodologia de leito fixo e expandido. O leito expandido possibilita a redução nos custos do processo de purificação, eliminando etapas de separação necessárias quando o extrato apresenta material em suspensão, que é o caso dos lactosoros. Nos ensaios realizados foram estudados as melhores condições de adsorção da a-Iactalbumina visando a sua purificação empregando adsorventes de troca iônica e de interação hidrofóbica. Também foram realizados ensaios em sistemas de tanque agitados para a determinação das isotermas e cinéticas de adsorção. Neste trabalho obteve-se a a-Iactalbumina com uma pureza acima de 80% e apresentando um fator de purificação de 5 vezes utilizando as resinas de interação hidrofóbica com única etapa de purificação Abstract: The interest and applications of biotechnology products has been increasing the development of new recovery and purification processes for proteins. The bovine milk serum, obtained from casein manufacture for cheese production, is mostly rejected into watercourse, causing problems to the environment due to its high biological oxygen demand (BOD). The proteins of milk serum have excellent nutritious and pharmaceutical value, h oweve r, its application for protein enrichment of food products is limited due to its low content in the milk serum. The main proteins of milk serum are: ~-Iactoglobulins, a-Iactalbumine, bovine serum albumine, immunoglobulins, lactoperoxidase, lactoferrin, lisozime and other lower content proteins which have a high aggregate value. The a-Iactalbumine acts in the human organism by stimulating the agents of the immunologycal system due to increasing on glutathione levei in several organs and blood, resulting in benefits for patients of some diseases like Parkinson and Alzheimer's iII, cancer and AIDS. It was studied in this work the separation processes of a-Iactalbumine, by chromatographic techniques making use of fixed and expanded bed methods. The expanded bed enables cost reduction on purification process by reducing separation steps used for removing suspended solids, as in case of milk serum extracts. In our experiments were studied the adsorption conditions of alactalbumine aiming at its purification by using ionic exchange and hydrophobic interaction adsorbents. Other experiments were accomplished at stirred tank systems for the determination of isotherms and adsorption kinetics. It was obtained, in this work, an a-Iactalbumine purity higher than 80%, with a five fold purification factor by using the hydrophobic interaction resins in a single purification ste Mestrado Desenvolvimento de Processos Biotecnológicos Mestre em Engenharia Química

O presente trabalho teve como objetivo desenvolver uma bebida lactea fermentada desenvolvida com soro de leite, extrato de erva-mate e estevia e avaliar as caracteristicas fisico-ligadas, microbiologicas e sensoriais do produto. Primeiramente, nas formulacoes da bebida, variou-se a concentracao de soro / leite (25/75; 50/50 e 75/25) e de extrato de erva-mate (0,1; 0,25 e 0,35%, m / v) e mantendo-se fixas como regras de stevia (0,75%, m / v), fermento lacteo (3,5%, m / v), sorbato de potassio (0,1%, m / v) e cacau em po (1%, m / v). As formulacoes foram caracterizadas fisico-quimicamente quanto ao pH, acidez, lactose, proteina, gordura, minerais totais, cor (L, a * eb *), sinerese, microbiologica e aceitabilidade. Os resultados das analises fisico-aglomerados demostram uma variacao entre as devedor, a qual pode estar relacionada com a quantidade de soro / leite, extrato de erva-mate e stevia adicionadas. A partir dos resultados da aceitabilidade das 7 formulacoes, realizou-se novos ensaios com a combinacao fixa de soro / leite (50/50), extrato de erva-mate (0,25%, m / v) fermento lacteo (3,5 %, m / v), sorbato de potassio (0,1%, m / v) e cacau em po (1%, m / v) e variacao na concentracao de estevia (0,625; 0,750; 0,875%, p / v) , realizando-se novamente analises fisicas-compostas, microbiologicas e sensorial. A composto com 50/50% de soro / leite, 0,25% de extrato de erva-mate, 3,5% de fermento lacteo, 0,1% de sorbato de potassio, 1% de cacau em po, 0,25 % de extrato de erva-mate e 0,650% de estevia apresentada um indice de aceitacao de 81,57%. Este indice de aceitacao demostra que uma bebida lactea fermentada carregada com adicao de soro de leite

Volatile sulfur compounds in ultra-pasteurized (UP) milk are the major contributors to sulfur/burnt and eggy flavors, and these flavors are disliked by consumers. Previous research has established distinct differences in flavor profiles of fluid milk processed by high temperature, short time pasteurization (HTST) and UP by direct steam injection (DSI-UP) or indirect heat (IND-UP). An understanding of the contribution of the individual milk proteins to sulfur off-flavors would clarify the source of sulfur flavors in UP milks. The objective of this study was to determine the source of volatile sulfur compounds in fluid milk with a specific focus on the comparison of heat treatment effects on milks by HTST and UP. Formulated skim milks (FSM) were manufactured by blending micellar casein concentrate and serum protein isolate (SPI). Three different caseins as a percentage of true protein (FSM95, FSM80, and FSM60) were formulated to determine the source of sulfur/burnt and eggy flavors. Freshly processed micellar casein concentrate or SPI were blended to achieve a true protein content of about 3.2%. Raw skim milk served as a control. Skim milk and FSM were pasteurized at 78°C for 15 s (HTST) or 140°C for 2.3 s by IND-UP or DSI-UP. The experiment was replicated twice. Sensory properties of milks and FSM were documented by descriptive sensory analysis. Volatile sulfur compounds in milks and FSM were evaluated using solid-phase microextraction followed by gas chromatography-triple quadrupole mass spectrometry combined with a sulfur selective flame photometric detector. The FSM with higher SPI as a percent of true protein had higher sensory sulfur/burnt and eggy flavors along with elevated concentrations of hydrogen sulfide and carbon disulfide compared with skim milk or FSM with lower proportions of SPI. Sulfur compounds including dimethyl sulfide, dimethyl disulfide, dimethyl trisulfide, dimethyl sulfoxide, and methional were not associated with sulfur/burnt and eggy flavors, which suggests that these compounds may not specifically contribute to the sulfur/burnt and eggy off-flavors of UP milks. Sensory panelists found higher overall aroma impact, and cooked, sulfur/burnt, and eggy flavors for DSI-UP, followed by IND-UP and HTST. The combination of sensory and instrumental methods used in the current study effectively identified that milk serum proteins are the source of sulfur compounds in milk, and further confirmed the contribution of hydrogen sulfide and carbon disulfide to eggy and sulfur/burnt flavors, respectively.

Horns are living tissue and cows can use their horns for thermoregulatory purposes. We investigated the effect of the presence of horns on the metabolome of milk serum and lipidome of milk fat, to assess the physiological effect of dehorning. Milk sampling took place at low ambient temperatures of −6 to 2°C. Horned and dehorned cows were kept in a mixed herd of Holstein Friesian and Brown Swiss cows. The hypothesis was that horned cows needed to increase their metabolism to compensate for additional heat loss through the presence of their horns. No differences were observed in milk yield, milk solids, and somatic cell counts between horned and dehorned cows. For the milk metabolome, horned cows showed an upregulation of several glucogenic AA that could be transformed into glucose for energy supply and a downregulation of sugar intermediates and γ-glutamylcysteine compared with dehorned cows. The fatty acid (FA) composition in horned cows showed a shift toward decreased odd medium-chain FA (C7:0, C9:0, and C11:0) and increased cis-vaccenic acid (C18:1n-7 cis-11) and stearidonic acid (C18:4n-3). The changes in milk composition related to additional heat loss in horned cows indicate a competition in C3 metabolism for glucose synthesis and de novo FA synthesis under cold stress.

Orientador: Aloisio Jose Antunes Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos Resumo: As proteínas da clara do ovo de galinha têm sido extensivamente usadas como ingredientes em alimentos processados, devido a suas propriedades funcionais, tais como gelatinização e formação de espuma. Durante o armazenamento, podem ocorrer mudanças nas características físicas, microbiológicas, químicas e funcionais dos ovos, que dependem das condições de armazenamento. As vantagens de utilizar filmes e coberturas comestíveis podem ser justificadas pela manutenção das propriedades funcionais dos alimentos, através da diminuição da perda de umidade e da diminuição do transporte de gases (02 e CO2). A aplicação de cobertura em ovos com casca reduz a perda de peso e mantém a qualidade interna do produto. Os objetivos foram quantificar a perda de peso, os valores da unidade Haugh, o pH e os teores de s-ovalbumina do albume, além de determinar alterações na estabilidade da espuma da clara, no perfil de textura e na umidade espremível de géis da clara de ovos frescos e de ovos com e sem cobertura à base de concentrado protéico de soro de leite, armazenados a 25°C, por 3,7, 10, 14,21 e 28 dias. Durante todo o período de estocagem houve decréscimo dos valores de unidade Haugh e perda de peso, tanto para os ovos com cobertura como para os ovos sem cobertura. Já o pH do albume aumentou. A variação dos valores nos ovos com cobertura foi menor do que para os ovos sem cobertura. Os valores da unidade Haugh para ovos com cobertura foram semelhantes aos de ovos armazenados sob refrigeração, encontrados em referências bibliográficas, no mesmo período de armazenamento. O volume de líquido drenado foi maior (P < 0,01) em ovos sem cobertura do que em ovos com cobertura armazenados a 25°C, em todos os períodos de armazenamento. A diferença no terceiro dia de armazenamento foi de 59% entre os ovos com cobertura e ovos sem cobertura, e no vigésimo oitavo dia foi 202%. Durante o período de armazenamento, ocorreu um aumento do pH e do volume do líquido drenado nos ovos sem cobertura. O teor de s-ovalbumina foi maior em ovos sem cobertura, comparados aos ovos com cobertura, em todos os períodos de armazenamento (P

The culture of rice has great potential to constitute the production systems of annual perennial crops, mainly in small properties. This allows this system to integrate with other activities such as the dairy industry, taking advantage of the milk by-product, reducing the environmental liability for disposal in the environment. Therefore, the objective was to evaluate the growth and productivity of rice grains under different doses of acid milk serum (0 m3 ha-1, 5 m3 ha-1, 10 m3 ha-1, 20 m3 ha-1, 50 m3 ha-1, 100 m3 ha-1, and 150 m3 ha-1), in soil applications with monitoring of fertility and electrical conductivity levels. The rice was sown in the UFLA experimental field, using the BRSMG Esmeralda lineage. Seven different doses were used, divided into two applications via soil. No cover fertilization was carried out. Yield and crop growth variables were determined by monitoring the nutrient content in the soil profile and the sodification and salinization capacity of the residue. It was found that the increase in serum doses increased the electrical conductivity and the percentage of exchangeable sodium by 1.75 and 7.75 compared to the control, but did not cause the process of sodification and salinization of the soil. As acid milk serum is a significant source of potassium, its use in the soil did not promote significant grain yield. It was concluded that the acid milk serum in the doses used did not cause sodification or salinization of the soil. This residue is the safe dose that can be used as potassium and nitrogen. The acid milk serum in the tested conditions did not increase the growth of the culture.

To establish the effect of the presence of milk serum proteins on heat-induced changes to lactoferrin, lactoferrin alone, and lactoferrin mixed with either milk serum or β-lactoglobulin was heated at 65 °C, 70 °C and 75 °C for 30 min. After heating, the effect of milk serum proteins on aggregation of lactoferrin was characterized, after which the effect of such aggregation on digestion and bacteriostatic capacity of lactoferrin were determined. The presence of milk serum proteins accelerated the aggregation of lactoferrin during heating through thiol/disulphide interchange. Lactoferrin also formed disulphide-linked aggregates when it was heated with β-lactoglobulin. Protein aggregates formed at 75 °C were much more resistant to infant digestion, causing decreased peptide release from lactoferrin. Heating lactoferrin and milk serum proteins together accelerated the loss of bacteriostatic activity upon heating. In conclusion, heat-induced aggregation of lactoferrin with milk serum proteins affected both its digestion and its bacteriostatic activity.

In bovine milk serum, the whey proteins with the highest N-glycan contribution are lactoferrin, IgG, and glycosylation-dependent cellular adhesion molecule 1 (GlyCAM-1); GlyCAM-1 is the dominant N-linked glycoprotein in bovine whey protein products. Whey proteins are base ingredients in a range of food products, including infant formulas. Glycan monosaccharide composition and variation thereof may affect functionality, such as the interaction of glycans with the immune system via recognition receptors. It is therefore highly relevant to understand whether and how the glycosylation of whey proteins (and their functionality) can be modulated. We recently showed that the glycoprofile of GlyCAM-1 varies between cows and during early lactation, whereas the glycoprofile of lactoferrin was highly constant. In the current study, we evaluated intercow differences and the effects of macronutrient supply on the N-linked glycosylation profiles of the major whey proteins in milk samples of Holstein-Friesian cows. Overall, approximately 60% of the N-glycan pool in milk protein was sialylated, or fucosylated, or both; GlyCAM-1 contributed approximately 78% of the total number of glycans in the overall whey protein N-linked glycan pool. The degree of fucosylation ranged from 44.8 to 73.3% between cows, and this variation was mainly attributed to the glycans of GlyCAM-1. Dietary supplementation with fat or protein did not influence the overall milk serum glycoprofile. Postruminal infusion of palm olein, glucose, and essential AA resulted in shifts in the degree of GlyCAM-1 fucosylation within individual cows, ranging in some cases from 50 to 71% difference in degree of fucosylation, regardless of treatment. Overall, these data demonstrate that the glycosylation, and particularly fucosylation, of GlyCAM-1 was variable, although these shifts appear to be related more to individual cow variation than to nutrient supply. To our knowledge, this is the first report of variation in glycosylation of a milk glycoprotein in mature, noncolostral milk. The functional implications of variable GlyCAM-1 fucosylation remain to be investigated.

A fast and reliable method for the identification of milk from different mammalians was developed by using 31P NMR metabolite profile of milk serum coupled to multivariate analysis (PCA and classification models UNEQ, SIMCA and K-NN). Ten milk samples from six different mammalians, relevant to human nutrition (human, cow, donkey, mare, goat, sheep), were analyzed and eight monophosphorylated components were identified and quantified: phosphocreatine (PCr), glycerophosphorylcholine (GPC), glycerophosphorylethanolamine (GPE), N-acetylglucosamine-1-phosphate (NAcGlu-1P), lactose-1-phosphate (Lac-1P), galactose-1-phosphate (Gal-1P), phosphorylcholine (PC), glucose-6-phosphate (Glu-6P). PCA showed interesting clustering based on the animal genus. K-NN can be successfully used to discriminate between donkey and cow samples while UNEQ class-modeling resulted more suitable for compliance verification. Results confirm the natural variability of milk samples among different species. These data highlight the great potentials of NMR/multivariate analysis combined method in the rapid analysis of phosphorylated milk serum metabolites for milk origin assessment and milk adulteration detection., Graphical abstract Image 1, Highlights • A31P NMR and multivariate protocol for analysis of milk metabolites is proposed. • Samples from different mammalians relevant to human nutrition were analyzed. • Eight monophosphorylated components were identified and quantified. • Multivariate analysis showed interesting clustering based on the animal genus. • Results confirm the natural variability of milk samples among different species.

The turbidity of milk prohibits the use of optical density measurements for strain characterizations. This often limits research to laboratory media. Here, we cleared milk through centrifugation to remove insoluble milk solids. This resulted in a clear liquid phase, termed milk serum, in which optical density measurements can be used to track microbial growth until a pH of 5.2 is reached. At pH 5.2 coagulation of the soluble protein occurs, making the medium opaque again. We found that behavior in milk serum was predictive of that in milk for 39 Lactococcus lactis (R2 = 0.81) and to a lesser extent for 42 Lactiplantibacillus plantarum (formerly Lactobacillus plantarum; R2 = 0.49) strains. Hence, milk serum can be used as an optically clear alternative to milk for comparison of microbial growth and metabolic characteristics. Characterization of the growth rate, specific acidification rate for optical density at a wavelength of 600 nm, and the amount of acid produced per unit of biomass for all these strains in milk serum, showed that almost all strains could grow in milk, with higher specific acidification and growth rates of Lc. lactis strains compared with Lb. plantarum strains. Nondairy Lc. lactis isolates had a lower growth and specific acidification rate than dairy isolates. The amount of acid produced per unit biomass was relatively high and similar for Lc. lactis dairy and nondairy isolates, as opposed to Lb. plantarum isolates. Lactococcus lactis ssp. lactis showed slightly lower growth and acidification rates when compared with ssp. cremoris. For Lc. lactis strains a doubling of the specific acidification rate occurred with a doubling of the maximum growth rate. This relation was not found for Lb. plantarum strains, where the acidification rate remained relatively constant across 39 strains with growth rates ranging from 0.2 h−1 to 0.3 h−1. We conclude that milk serum is a valuable alternative to milk for high-throughput strain characterization during milk fermentation.

The use of antibiotics to prevent bovine mastitis is responsible for the emergence and selection of resistant strains. Lactic acid bacteria (LAB) could be introduced into animal feed as an alternative prevention method that would bypass the risk of resistance development. In previous research, we demonstrated that two probiotic LAB strains isolated from bovine milk were capable of stimulating the production of antibodies and the host’s immune cellular response in the udder. The present study aimed to elucidate whether the antibodies of animals inoculated with these strains were able to increase phagocytosis by neutrophils and inhibit the growth of different mastitis-causing pathogens. Moreover, the effect of LAB on the expression of pro-inflammatory cytokines was assessed. Ten animals were inoculated intramammarily with 106 cells of the two strains at dry-off. The blood serum was tested for its ability to opsonize bovine mastitis pathogens, the in vitro bactericidal activity of bovine blood and milk against these pathogens was determined, and cytokine mRNA expression was quantified in milk somatic cells. The inoculated animals did not show abnormal signs of sensitivity to the LAB. Their blood serum significantly enhanced the phagocytosis of Staphylococcus spp. and the LAB. Escherichia coli and Streptococcus uberis were inhibited by the milk serum but not the blood serum, whereas Staphylococcus aureus and Staphylococcus haemolyticus were inhibited by both. In regard to cytokine expression, interleukin (IL)-1β increased markedly for up to 4 h post-inoculation, and an increase in IL-8 was observed 4, 12 and 24 h after inoculation. Tumour necrosis factor-α mRNA increased 1 and 2 h after inoculation and a significant difference was registered at 6 h for interferon-γ. This rapid immunomodulatory response shows that inoculating animals with LAB at dry-off, when they are especially susceptible, could be a useful strategy for the prevention of bovine mastitis.

The aim of this study was characterization of some dairy drinks based on Milk Serum regarding major whey proteins (WP) and free amino acids (FAAs) using reversed phase high performance liquid chromatographic (RP-HPLC) methods. The studied WP, -lactalbumin (-La), bovine serum albumin (BSA), -lactoglobulin A (-Lg A) and -lactoglobulin B (-Lg B) were separated on Aeris XB-C18 column at 214 nm detection. The RP-HPLC method was validated by selectivity, linearity (R2 ≥0.99), sensitivity (LOQ, 1.35–10.08 µg mL−1), accuracy (recovery 96.79-103.07%) and precision (% RSD ≤ 4.13%). The total studied WP in studied dairy drinks varied between 1.42 and 3.047 g·L-1. The chromatographic profile of FAAs (aspartic acid, glutamic acid, serine, histidine, arginine, glycine, threonine, alanine, tyrosine, cysteine, tryptophan, methionine, valine, phenylalanine, isoleucine, leucine and lysine) was determined in lyophilized concentrate of Milk Serum by RP-HPLC using pre-column derivatization reaction with orthophthalaldehyde (OPA). The total studied FAAs in studied samples varied between 1.103 and 1.119 mg·g-1. Moreover, the Milk Serum showed bacteriostatic activity against two bacterial strains Escherichia coli and Staphylococcus aureus. The obtained results confirm that dairy drinks based on the Milk Serum constitutes a valuable sources of bioactive components with benefits for human healthy nutrition.

This study provided insights into the degradation of human milk proteins in an in vitro infant digestion model by comparing colostrum (week 1) and mature milk (week 4) of 7 Chinese mothers individually. In this study, we adapted the exiting INFOGEST in vitro model, to conditions representative to infants (0 to 3 month-old). The level of undigested proteins was analyzed by LC-MS/MS after gel-electrophoretic separation and in-gel digestion. The BCA protein assay showed that the total undigested milk protein content decreased from the start to the end of digestion with variations between mothers, especially in the gastric phase (25–80%). Undigested proteins could also still be found after the intestinal phase, ranging from 0.5 to 4.2% of initial protein content. Based on LC-MS/MS analysis, milk protein digestion varied between the mothers individually, especially during the gastric phase. No differences could be observed between protein digestion from colostrum and mature milk after the intestinal phase. The highest levels of proteins remaining after intestinal digestion can be linked to the group immune-active proteins, for all mothers. The level of protease inhibitors and total protein content in the milk did not correlate with the overall proteolysis during digestion. The results also showed that milk serum proteins partly remained after the gastric phase, with 33% remaining from colostrum and 37% remaining from mature milk. More than 40 milk serum proteins were detected after the intestinal phase. Some of the highly abundant milk serum proteins (lactoferrin, serum albumin, bile salt-activated lipase, immunoglobulins, α1-antichymotrypsin) were still partially present intact after the intestinal phase, for all mothers. Caseins were also not completely digested in the gastric phase, with 35% remaining from colostrum and 13% remaining from mature milk. Caseins, on the other hand, were almost completely digested after the intestinal phase. The complete degradation of caseins into peptides might be related to their structural features. Overall, this study showed that digestion differed for the various human milk proteins by adapting an in vitro digestion model to infant physiological conditions, with the main differences between digestion of the milk from individual mothers being observed after gastric digestion.

Hepatitis E virus (HEV) infection is endemic in developing and developed countries. HEV was reported to be excreted in the milk of ruminants, raising the possibility of transmission of HEV infection through the ingestion of contaminated milk. Therefore, the detection of HEV markers in milk samples becomes pivotal. However, milk includes inhibitory components that affect HEV detection assays. Previously it was reported that dilution of milk matrix improves the performance of HEV molecular assay, however, the dilution of milk samples is not the best strategy especially when the contaminated milk sample has a low HEV load. Therefore, the objective of this study is to compare the effect of extraction procedures on the efficiency of HEV RNA detection in undiluted milk samples. In addition, we assessed the effect of the removal of milk components such as fats and casein on the performance of the molecular and serological assays of HEV. Phosphate buffered saline (PBS) and different milk matrices (such as whole milk, skim milk, and milk serum) were inoculated with different HEV inoculums and subjected to two different extraction procedures. Method A includes manual extraction using spin column-based extraction, while method B includes silica-based automated extraction. Method A was more sensitive than method B in the whole milk and skim milk matrices with a LoD95% of 300 IU/mL, and virus recovery yield of 47%. While the sensitivity and performance of method B were significantly improved using the milk serum matrix, with LoD95% of 96 IU/mL. Interestingly, retesting HEV positive milk samples using the high sensitivity assay based on method B extraction and milk serum matrix increased the HEV RNA detection rate to 2-fold. Additionally, the performance of HEV serological assays such as anti-HEV IgG and HEV Ag in the milk samples was improved after the removal of the fat globules from the milk matrix. In conclusion, HEV RNA assay is affected by the components of milk and the extraction procedure. Removal of inhibitory substances, such as fat and casein from the milk sample increased the performance of HEV molecular and serological assays which will be suitable for the low load HEV milk with no further dilutions.

Liquid milk protein concentrate with different beneficial values was prepared by membrane filtration and enzymatic modification of proteins in a sequential way. In the first step, milk protein concentrate was produced from ultra-heat-treated skimmed milk by removing milk serum as permeate. A tubular ceramic-made membrane with filtration area 5 &times, 10&minus, 3 m2 and pore size 5 nm, placed in a cross-flow membrane house, was adopted. Superior operational strategy in filtration process was herein: trans-membrane pressure 3 bar, retention flow rate 100 L&middot, h&minus, 1, and implementation of a static turbulence promoter within the tubular membrane. Milk with concentrated proteins from retentate side was treated with the different concentrations of trypsin, ranging from 0.008&ndash, 0.064 g&middot, L&minus, 1 in individual batch-mode operations at temperature 40 &deg, C for 10 min. Subsequently, inactivation of trypsin in reaction was done at a temperature of 70 &deg, C for 30 min of incubation. Antioxidant capacity in enzyme-treated liquid milk protein concentrate was measured with the Ferric reducing ability of plasma assay. The reduction of angiotensin converting enzyme activity by enzyme-treated liquid milk protein concentrate was measured with substrate (Abz-FRK(Dnp)-P) and recombinant angiotensin converting enzyme. The antibacterial activity of enzyme-treated liquid milk protein concentrate towards Bacillus cereus and Staphylococcus aureus was tested. Antioxidant capacity, anti-angiotensin converting enzyme activity, and antibacterial activity were increased with the increase of trypsin concentration in proteolytic reaction. Immune-reactive proteins in enzyme-treated liquid milk protein concentrate were identified with clinically proved milk positive pooled human serum and peroxidase-labelled anti-human Immunoglobulin E. The reduction of allergenicity in milk protein concentrate was enzyme dose-dependent.

Human milk contains, among others, a range of biofunctional components, like serum proteins and human milk oligosaccharides (HMOs). This PhD thesis provides information on the variability in serum proteins, serum protein N-glycans and HMOs in milk of individual mothers during lactation, and offers insights on the degradation of proteins using a newly developed infant (0–3 months) in vitro digestion model.The 15 most abundant HMOs cover >95% of the total HMO content. The mother’s secretor (Se) and Lewis (Le) histo-blood groups are related to specific fucosylated HMOs. Based on the total neutral fucosylated HMO concentrations in both Chinese and Dutch human milk, for the first time, Se+Le+ subgroups were identified. Variation among mothers was found in the composition of serum proteins in both colostrum and mature milk, although the group of immune-active proteins, enzymes, and transport proteins were the most abundant for all mothers. These three protein groups encompass many of the 15 most abundant proteins, covering >95% of the total protein concentrations, in both the Chinese and Dutch milk serum proteome. The serum protein N-glycan composition in human milk from 2 different lactational periods was investigated. Analysis showed that neutral fucosylated and nonfucosylated N-glycans dominated the human milk serum glycoproteome. Based on the levels of the individual N-glycans, for the first time, a clear difference can be observed between the milk of secretor and nonsecretor mothers. It also indicated that specific fucosylated N-glycans can be synthesized in the gland of nonsecretor mothers and even in much higher concentrations compared to secretor mothers. In an in vitro infant digestion model, the total milk protein content decreased from the start to the end of infant in vitro digestion with large variation between mothers, especially in the gastric phase (remaining between 25–80%). After intestinal digestion, still some undigested proteins could be found, ranging from 0.5% to 4.2% of the initial protein content, although no differences could be observed between colostrum and mature milk. More than 40 serum proteins could be detected after intestinal digestion. Overall, caseins are digested more than most serum proteins during digestion. Especially immune-active serum proteins, which are digested to a lower extent, might protect infants from pathogens.

Human milk provides a range of nutrients and bioactive components, which can support the growth and development of infants. However, human milk composition may change due to geographic and ethnic variation. This study investigated the variation of the Chinese human milk serum proteome based on mothers with different ethnicities living in different parts of China, using TMT labeling combined with Nano-LC Q Exactive HF MS/MS proteomics. In total, 693 proteins were identified and quantified in human milk serum from Yunnan (Han and Bai ethnicity), Gansu (Han and Tibetan ethnicity), Xinjiang (Uygur ethnicity), and Inner Mongolia (Mongolian ethnicity). The biological function distribution of identified proteins and the summed intensity of proteins belonging to each biological function were similar among groups. The five relatively highly abundant milk serum proteins, lactoferrin, serum albumin, polymeric immunoglobulin receptor, macrophage mannose receptor 1, and bile salt-activated lipase were not significantly different among different geographies and ethnicities. On the other hand, we found 34 proteins that did significantly differ with geography and ethnicity. Those significantly different proteins have known strong interaction in inflammation response and regulation of multi-organism processes. Taken together, biological function distribution was similar on both the qualitative and quantitative levels, and proteins with similar abundance are important in providing basic nutrition and protection for infants, whereas the significantly different proteins may be important for the healthy development of infants from different locations and ethnicities.

Context: Candida, leukoplakia and oral squamous cell carcinoma. Aims: The aim of the present study has assessed the association of Candida albicans with normal control group, leukoplakia and oral squamous cell carcinoma lesions of the oral cavity by using cornmeal broth +5% milk and milk serum liquid culture media. Settings and Design: Thepresent study comprised of total 80 patients,which included, 30 patients of Leukoplakia, 30 patients of oral squamous cell carcinoma and normal control group comprised of 20 healthy individuals who were not having any relevant medical, dental and habit history. Subjects and Methods: Saliva and Biopsy was taken from clinically suspected leukoplakia and oral squamous cell carcinoma lesions for the confirmation of histopathological diagnosis. Saliva samples were inoculated for fungal growth in Sabouraud Dextrose Agar, and culture-positive samples had undergone for the germ tube test. Germ tube-positive samples were further taken for chlamydospore production in milk serum and cornmeal broth +5% milk media separately at 8 and 16 h. Statistical Analysis Used: Chi-square test, Fischer extract test. Results: In the normal control group, no fungus growth was found; however, leukoplakia and oral squamous cell carcinoma showed fungus growth with positive germ tube test and chlamydospore formation; the result also showed rapid and quantitatively more chlamydospore formation in cornmeal broth +5% milk in comparison to serum milk culture media. There was no growth of Candida in mild and moderate form of leukoplakia. Whereas in a severe form of leukoplakia, there was more quantitative chlamydospore growth in cornmeal broth +5% milk liquid media. Conclusions: In this study, we have tried to compare the efficacy of cornmeal broth +5% milk and milk serum for the identification of C. albicans. Both the culture media were able to promote the growth of chlamydospore in C. albicans. Among different grades of leukoplakia, the growth of C. albicans was seen in severe dysplastic patient only while mild and moderate dysplasia showed no Candidal growth.

The aim of this work is to study the ability of whey flavor precursors to form new volatile components under model conditions. The formation of new aromas is considered in the aspect of a two-stage process - enzymatic cleavage of the precursors and the subsequent Maillard reaction. The properties of whey components by their ability to form or modify aromatic descriptors at the stages of enzymatic cleavage of taste precursors are considered. It is proved that targeted enzymatic reactions with whey components lead to the formation of sweet, meat, cheese flavors at various stages of whey protein breakdown after interaction with carbohydrates. The influence of the depth of proteolysis on the subsequent formation of aroma from precursors is considered. It has been established that the breakdown products of milk sugar in whey do not play a significant role in the formation of aroma. Unlike proteolytic enzymes, the use of lactase (β-galactosidase) did not lead to a change in the aromatic characteristics of the modified serum. An increase in the concentration of lactulose, as a potential precursor of aroma in the sugar-amine reactions of serum, gives only a sweetish tint and does not significantly affect the change in aroma. The processes of aroma formation in modified whey have been consistently investigated, and the concept of flavored food foam based on it has been tested. It was shown that the addition of sodium chloride 5-10% to the lower residue after rectification of fermented milk whey significantly increases the stability of the aroma of the final product. It was found that the specific aroma of whey, limiting its use in food, can be modified by using new approaches to reactions with aroma precursors. We consider the fixation of the obtained aromas “soup mushroom”, “broth”, “cheese” on an oil basis for subsequent concentration and use in various products.

Infant formula is used as a supplement for newborns. Although bovine milk-based infant formulas dominate the market, caprine milk-based infant formula has attracted increasing attention because of its lower allergenicity. This study compared the digestive peptidome of bovine and caprine milk serum proteins by using in vitro infant simulating conditions. The result showed that the degradation pattern of milk proteins was similar, whereas the digestive rates of milk proteins differed between bovine and caprine milks. Several proteins, such as α-lactalbumin (LALBA), β-lactoglobulin (LGB), serum amyloid A protein (SAA1), glycosylation-dependent cell adhesion molecule 1 (GLYCAM1), and lactotransferrin (LTF), released more peptides during digestion of caprine milk serum than during digestion of bovine milk serum; however, more peptides derived from αS1-casein (CSN1S1) were found in bovine digesta. In addition, antimicrobial-related peptides were mostly only found in caprine intestinal digesta. The results of this study may be useful in understanding the digestion characteristics of milk serum proteins and providing guidance on the improvement of infant formula.

Our previous study identified peaks in the 31 P nuclear magnetic resonance ( 31 P NMR) spectra of skim milk, denoting the interaction of different phosphate species such as inorganic and casein-associated phosphate during the separation of colloidal and serum phases of skim milk by microfiltration (MF) and diafiltration (DF). In the current study, we investigated the same samples generated by the aforementioned separation using attenuated total reflectance (ATR) Fourier transform infrared (FTIR) spectroscopy analysis. The results confirmed that the technique was not only capable of differentiating between the mineral equilibrium of the casein phosphate nanocluster (CPN) and milk serum, but also complemented the application of 31 P NMR. An ATR-FTIR broad band in the region of 1,055 to 1,036 cm −1 and a specific band at 1,076 cm −1 were identified as sensitive to the repartitioning of different phosphate species in milk in accordance with the 31 P NMR signals representing casein-associated phosphate and inorganic phosphate in the serum. A third ATR-FTIR signal at 1,034 cm −1 in milk, representing precipitated inorganic calcium phosphate, had not previously been detected by 31 P NMR. Thus, the results indicate that a combination of ATR-FTIR and 31 P NMR spectroscopies may be optimally used to follow mineral and protein phase changes in milk during membrane processing.

The article discusses the best options for the preparation of blood and milk samples for the purpose of diagnosing retroviral enzootic bovine leukosis in cattle and assessing the origin of dairy products from sick or healthy animals. The effectiveness of using samples in the native state and after their processing with the removal of red blood cells, their membranes, the fat component and casein were analyzed. It has been established that when performing screening analysis it can be carried out using whole blood and milk samples. This is especially possible in the case of diagnosis of the disease at the stage of its progressive development. The use of such samples significantly reduces analysis time and makes it simpler. But if it is necessary to carry out data verification and detection of cattle retroviral leukemia virus already in the early periods, then, in accordance with the results obtained by us, samples prepared in the form of blood serum or milk serum should be used. The latter should also be used to accurately assess the origin of milk, especially when it is necessary to control its quality when collecting from several animals, among which may be sick individuals, as well as in the early stages of the development of the pathological process.

This study investigated milk serum protein concentrate/lactose systems with varying ratios and how the morphology of the spray-dried particles of these systems could be described by the surface properties of the feed as well as the protein surface coverage of the particles. An extrapolation of the surface pressure of the feed to 0.3 s, the approximate time for molecular diffusion in an atomised droplet in the spray-dryer, showed a relationship with the particle morphology. At low protein concentrations (

In present study, 198, 169, 213 and 128 proteins were identified and quantified in human, cow, goat and yak milk serum respectively by using proteomics techniques. Large variations were observed between human and ruminant milk proteins. Human milk contained higher concentration of mucosal immune response, complement proteins and regulators. The concentration of bactericidal proteins were relatively higher in ruminants milk. Human milk exclusively expressed proteins important for delivery or utilization of nutrients. Peptidase inhibitors prevent the bioactive proteins/peptides in human milk from gastral-intestinal digestion. Protein composition among ruminants milk was similar but with variations. Goat milk contained high level of complement proteins but low level of corresponding regulators. In addition, the acute phase proteins were significantly higher in goat milk. Of note, osteopontin in yak milk was enriched, which could offer an alternative source for producing osteopontin and revealed possible nutritional value of yak milk.

This study investigated the effects of heating skim milk with soluble calcium salts, calcium chloride, calcium lactate, calcium gluconate and calcium lactobionate, on the physico-chemical and rheological properties of milk. Regardless of the type of salt added, the amount of casein in milk serum decreased and the amount of calcium in centrifuge sediment increased along with the serum calcium. The amount of calcium salt required for gelation and the gel firmness (G') varied depending on the salt. The aCa2+ order in milk was calcium chloride>calcium lactate>calcium gluconate>calcium lactobionate. With the same amount of added calcium salt, the final G' of milk gel followed the same order. The findings from this study suggest that the different association constants and formation of an intermediate calcium complex with hydroxycarboylate groups affected the calcium ion activity, which may influence the gelation properties in milk.

This study aims to evaluate the in vivo efficacy in terms of elasticity, hydration and anti-wrinkle enhancement of a new cosmetic cream formulation: Apollinea anti-ageing facial cream. The latter is O/W (oil/water) emulsion with high volume percentages of snail secretion filtrate and donkey milk serum in its composition. The investigation has been performed on (twelve) healthy subjects: 8 (eight) females, and 4 (four) males, all presenting evident skin wrinkles in typical facial areas (e.g. forehead and eye contour). The subjects had to apply about 2 grams of Apollinea anti-ageing facial cream every day. The efficacy of the new cosmetic formulation was assessed before and after the cosmetic treatment, precisely 2 hours, to assess the short-term efficacy, and 40 days after, this to evaluate the product long-term efficacy. Elasticity enhancements have been tested through measurements of the viscoelastic properties of the skin, whereas the hydration parameter through measurements of the skin and subcutaneous tissues dielectric constant. All the latter measurements have been performed with specific instrumentation, as discussed more in detail in the following. To evaluate the anti-wrinkles efficacy instead, we proceeded with digital camera imaging and image analysis processed through ImageJ software. The new Apollinea facial cream cosmetic formulation, based on donkey milk whey and snail secretion filtrate from Helix Aspersa Muller, showed significant enhancement effects on skin elasticity and hydration as well as an important skin-wrinkles-reduction.

Whey is a low calorie milk serum remaining after casein removal in cheese manufacturing. Composition and properties of whey depend on the type of cheese production process, type and the quality of the used milk. Whey mostly consists of lactose, proteins, vitamins, minerals, enzymes, hormones and growth factors. Functional and nutritional roles of whey components meet the requirements for dietary food. Whey proteins are liquid fractions of 20% milk proteins, which remain in whey after casein removal. The increase in the temperature of milk processing leads to denaturation of proteins and a decrease in their content. The most common whey proteins are albumin and globulin. Separation and fractionation of whey proteins may be carried out as precipitation, combined with acid treatment, high temperature treatment or centrifugation (centry-whey process), and as a membrane process (usually ultra- or microfiltration followed by a diafiltration and spray drying). Whey proteins can also be separated based on their molecular mass, charge and hydrophobicity. It is a well-known fact that whey proteins possess admirable nutritional and functional components, such as essential amino acids, bioactive peptides, antioxidants and immunopotentiators. Research of bioactive compounds and nutrients pushed whey protein to the forefront of the functional food sector, therefore, the implementation of these proteins in the diet and supplements has the potential to address many metabolic imbalance-caused diseases, and it seems imperative to harness their potential.

One of the main factors determining physicochemical and structural properties of the dry permeate of spray drying is lactose which is present in permeate as a basic phase. If lactose is in the β-form in a dry powder, the powder easily absorbs moisture from the atmosphere during storage, causing agglutination, clumping, and caking of the product. Prevention of negative consequences associated with increased hygroscopicity, is ensured by the directed crystallization of lactose in a condensed product before spray drying. The purpose of this investigation was to study the crystallization process of lactose in permeate under industrial conditions when using various seed materials. Results of the studies showed that the type of a crystal forming agentdoes not affect the size and shape of lactose crystals and the level of lactose crystallization. Seed material is of critical significance as it will ensure both a high level of crystallization and homogenous crystals which, in general, leads to an increase in melting temperature, due to a decrease in the proportion of amorphous lactose, as well as improves the drying process.

Whey (milk serum) is produced by the dairy industry during the manufacture of cheese. In addition to being a valuable source of functional and nutritional proteins, whey also presents almost all the lactose from the original whole milk. However, many industries still consider the whey as an effluent, which can cause serious environmental problems when not properly treated. Therefore, it is important to develop alternatives for the adequate use of whey. The lactose obtained from whey permeate can be converted into lactulose, a prebiotic which may be metabolized in the intestine by probiotic bacteria, such as Lactobacillus sp. and Bifidobacterium sp., through enzymatic isomerization or by using alkaline catalysts, with minimal secondary reactions and high yield. This manuscript provides information about various techniques used to produce lactulose, its purification and analysis, as well as its mechanisms of action and alternative applications in food products and medicines.

Milk whey proteins are precursors of more than a hundred bioactive peptides. These peptides may be formed in the digestive process in the gastrointestinal tract, as well as under the influence of lactic acid bacteria proteases and milk-clotting preparations. Among them, inhibitors of angiotensin-converting enzyme, opioid and bactericidal peptides, immunomodulatory and hypocholesterolemic peptides, peptides that have an effect on intestinal motility, appetite, and others have been found. Bioactive peptides from whey proteins are of considerable interest for the production of functional products or products with prophylactic properties. The creation of such products includes the obtaining of bioactive peptides of define biological action. Taking into account the specifics of their placement in the primary structure, it is advisable to use homogeneous protein precursors from milk whey to obtain them. One of the methods for one-stage separation of whey proteins is disc-electrophoresis in the native PAGE system for alkaline and neutral proteins. The purpose of this work was to determine the parameters of the preparative variant of disc-electrophoresis for the selection of homogeneous whey proteins. The research was performed on a modified apparatus of Stadier type. Proteins of milk whey were isolated by the precipitation of caseins at an isoelectric point. The precipitate of caseins was separated by centrifugation. Homogenous α-lactalbumin (α-LA) and β-lactoglobulin (β-LG) fractions were used to identify milk whey proteins, which were obtained by double gel-filtration on Sephadex G-100 (fine). Homogeneity and fractional composition of milk serum proteins were determined by analytical disс-electrophoresis in plates of polyacrylamide gel. The concentration of proteins in the samples was determined spectrophotometrically (spectrophotometer SF-46) at a wavelength λ = 280 nm.The obtained results prove the feasibility of using PAGЕ plates for the preparative fractionation of serum milk proteins. The allowable duration and conditions for identification of protein fractions are established. It was shown that the yield of proteins in the composition of homogeneous fractions was 69.2%. In this case, the correlation between the number of obtained fractions (β-LG, α-LA, BSA and IG) is close to their distribution in milk serum.