Short communication: estimate of non electrostatic interaction free energy parameters for milk fat globules

The surface free energy components for bovine milk fat globules (MFG) were calculated by solving a modified Young-Good-Fowkes-Girafalco equation with experimentally obtained contact angles for liquids of known surface tension components. The surface free energy of MFG was dominated by the electron donor term (gamma-), which formed the basis of a significant repulsive acid-base interaction free energy term at the minimum equilibrium distance. Consideration of the decay of the acid-base and Lifshitz-van der Waals energy term over distance within the context of an "extended DLVO" model shows that physical contact of MFG in the absence of electrostatic repulsion (at their isoelectric point) is prohibited by the acid-base term but that "pseudo-flocculation" might occur at 4- to 5-nm separation in an energy minimum that was approximately 15 kT for MFG of diameter = 1.0 microm.     

Technical note: Characterizing individual milk fat globules with holographic video microscopy

We used in-line holography to create images of individual milk fat globules in diluted samples of milk. Analyzing these images with the exact Lorenz-Mie light scattering theory then yields the droplets’ radii with nanometer resolution and their refractive indexes to within one part in a thousand. This procedure rapidly and directly characterizes both the quantity and quality of fat in milk.     

Chemical and structural characterisation of almond oil bodies and bovine milk fat globules

Lipids in almonds are present as oil bodies in the nut. These oil bodies are surrounded by a membrane of proteins and phospholipids and are a delivery vehicle of energy in the form of triglycerides, similarly to the more studied bovine milk fat globule membrane. Chemical, physical and microscopic analyses revealed major differences in the composition and structure of almond oil bodies and bovine milk fat globules. The lipids of both natural emulsions differed in degree of unsaturation, chain length, and class. The almond oil body membrane does not contain any cholesterol or sphingomyelin unlike the bovine milk fat globule membrane. Therefore, the phospholipid distribution at the surface of the oil bodies did not present any liquid-ordered domains. The membranes, a monolayer around almond oil bodies and a trilayer around bovine fat globules, may affect the stability of the lipid droplets in a food matrix and the way the lipids are digested.     

Size distribution of fat globules in human colostrum, breast milk, and infant formula

Only a few results are available on the size of human milk fat globules (MFG), despite its significance regarding fat digestion in the infant, and no data are available at <24 h postpartum (PP). We measured the MFG size distribution in colostrum and transitional human milk in comparison with fat globules of mature milk and infant formula. Colostrum and transitional milk samples from 18 mothers were collected regularly during 4 d PP and compared with mature milk samples of 17 different mothers and 4 infant formulas. The size distribution was measured by laser light scattering. For further characterization, the ζ-potential of some mature MFG was measured by laser Doppler electrophoresis. The MFG diameter decreased sigmoidally in the first days. At <12 h PP, the mode diameter was 8.9 ± 1.0 μm vs 2.8 ±0.3 μm at 96 h PP. Thus, the surface area of MFG increased from 1.1 ±0.3 to 5.4 ±0.7 m²/g between colostrum and transitional milk. In mature milk, the MFG diameter was 4 μm on average and increased with advancing lactation, whereas the droplets in infant formula measured 0.4 μm. The ζ potential of mature MFG was −7.8 ± 0.1 mV. The fat globules are larger in early colostrum than in transitional and mature human milk and in contrast with the small-sized fat droplets in infant formula. Human MFG also have a low negative surface charge compared with bovine globules. These structural differences can be of nutritional significance for the infant.     

Native vs. damaged milk fat globules: membrane properties affect the viscoelasticity of milk gels

The storage modulus G' of rennet and acid milk gels filled with milk fat globules was measured as a function of the fat globule surface composition (native milk fat globule membrane, caseins and whey proteins, or a mixture of the three due to mechanical treatments) and surface area (i.e., the fat globule size). By different technological procedures, it was possible to obtain fat globules of constant surface composition but various sizes, and vice-versa, which had never been done. For both rennet and acid gels, a critical fraction of the fat globule surface covered by caseins and whey proteins was identified (approximately 40%), beyond which G' increased. Below this threshold, the gel viscoelasticity was unaffected by mechanical treatments. When the diameter of native milk fat globules decreased, the G' of rennet gels increased slightly, whereas that of acid gels decreased sharply. For both types of gels, G' increased when the diameter of partially disrupted fat globules decreased. For recombined globules completely covered with caseins and few whey proteins, G' increased with fat globule surface area for rennet gels whereas it decreased for acid gels. With the help of confocal microscopy and in the light of general structural differences between rennet and acid gels, a mechanism is proposed for the effect of fat globule damage and diameter on G', depending on the interactions the globules can undergo with the casein network.     

Membrane-rich milk fat diet provides protection against gastrointestinal leakiness in mice treated with lipopolysaccharide

Milk fat globule membrane is a protein-lipid complex that may strengthen the gut barrier. The main objective of this study was to assess the ability of a membrane-rich milk fat diet to promote the integrity of the gut barrier and to decrease systemic inflammation in lipopolysaccharide (LPS)-challenged mice. Animals were randomly assigned to one of 2 American Institute of Nutrition (AIN)-76A formulations differing only in fat source: control diet (corn oil) and milk fat diet (anhydrous milk fat with 10% milk fat globule membrane). Each diet contained 12% calories from fat. Mice were fed diets for 5 wk, then injected with vehicle or LPS (10 mg/kg of BW) and gavaged with dextran-fluorescein to assess gut barrier integrity. Serum was assayed for fluorescence 24 h after gavage, and 16 serum cytokines were measured to assess the inflammatory response. Gut permeability was 1.8-fold higher in LPS-challenged mice fed the control diet compared with the milk fat diet. Furthermore, mice fed the milk fat diet and injected with LPS had lower serum levels of IL-6, IL-10, IL-17, monocyte chemotactic protein (MCP)-1, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, and IL-3 compared with LPS-injected mice fed the control diet. The results indicate that the membrane-rich milk fat diet decreases the inflammatory response to a systemic LPS challenge compared with corn oil, and the effect coincides with decreased gut permeability.     

Symposium review: Fat globules in milk and their structural modifications during gastrointestinal digestion

The fat globules in milk are unique oil droplets that are stabilized by a specific and structurally complex membrane, the milk fat globule membrane (MFGM). In the last decade, excellent progress has been made on studying the structure of the milk fat globules and the MFGM and how common processing treatments affect these structures to deliver dairy products with improved functional properties. Although the digestion of milk fat to deliver energy and lipid-soluble nutrients is essential for survival of the neonate, there is little understanding of the complex processes involved. The structural alterations to fat globules during gastrointestinal processing affect the way in which milk fat is digested, absorbed, and metabolized. The packaging of these globules within the MFGM or in other forms may affect the bioaccessibility of raw or processed milk fat globules; in turn, this may affect access of the gastrointestinal enzymes to the globules and, therefore, may influence the rate and extent of lipid digestion. This review focuses on recent advances in understanding milk fat globules during gastrointestinal digestion, including the effects of processing on their bioavailability and the kinetics of lipid digestion. Possible effects of the dairy matrix on lipid digestion and physiological responses are briefly described.     

Analysis of major proteins and fat fractions associated with mare's milk fat globules

Although several studies have aimed to identify mare's milk proteins, only the major whey proteins and some caseins have yet been characterized. Incomplete sequencing of the equine genome and the difficulty of recovering highly hydrophobic proteins mean that little is known to date about the proteins associated with milk fat globules, which have been shown to play an important role in newborns' defense mechanisms. The fat fraction, in particular the distribution of unsaturated fatty acids, has been more extensively studied, but complex lipids are only partially elucidated. This study reports a 2-DE approach combined with a powerful method for de novo protein sequencing, and quali-quantitative data on complex lipid composition determined by high performance TLC (HPTLC) and GC. The presence in mare's milk of long-chain highly unsaturated fatty acids, and the evidence of close similarity between equine and human milk fat globule membrane proteins, support the use of mare's milk for human nutrition.     

人乳脂肪球的研究进展

人乳是婴儿最理想的食物,人乳脂肪是人乳中的重要组成,不仅是婴儿生长发育所需能量的主要来源,也提供了各种脂溶性营养物质。人乳脂肪在乳清相中并非呈完全均匀地分散,而是以人乳脂肪球的形态稳定存在。人乳脂肪球是由一个三层的膜结构包裹住中心层状排列的甘油三酯组成,这种结构不仅维持了乳液本身的稳定性,还为婴儿的生长发育提供了特有的营养学功能。近年来,这种天然而特殊的物理形态引起了学者们的广泛关注。对国内外人乳脂肪球的相关文献进行综述,主要包括人乳脂肪球的形成、组成、营养学意义以及其在加工过程中的变化,并对今后研究的方向和重点进行了展望。     

Proteolysis of milk fat globule membrane proteins during in vitro gastric digestion of milk

The influence of gastric proteolysis on the physicochemical characteristics of milk fat globules and the proteins of the milk fat globule membrane (MFGM) in raw milk and cream was examined in vitro in simulated gastric fluid (SGF) containing various pepsin concentrations at pH 1.6 for up to 2 h. Apparent flocculation of the milk fat globules occurred in raw milk samples incubated in SGF containing pepsin, but no coalescence was observed in either raw milk samples or cream samples. The changes in the particle size of the fat globules as a result of the flocculation were dependent on the pepsin concentration. Correspondingly, the physical characteristics of the fat globules and the composition of the MFGM proteins in raw milk changed during incubation in SGF containing pepsin. The major MFGM proteins were hydrolyzed at different rates by the pepsin in the SGF; butyrophilin was more resistant than xanthine oxidase, PAS 6, or PAS 7. Peptides with various molecular weights, which altered with the time of incubation and the pepsin concentration, were present at the surfaces of the fat globules.     

Identification of ABCA1 and ABCG1 in milk fat globules and mammary cells--implications for milk cholesterol secretion

The ATP-binding cassette (ABC) transporters ABCA1 and ABCG1 play an important role in cellular cholesterol homeostasis, but their function in mammary gland (MG) tissue remains elusive. A bovine MG model that allows repeated MG sampling in identical animals at different functional stages was used to test whether 1) ABCA1 and ABCG1 protein expression and subcellular localization in mammary epithelial cells (MEC) change during the pregnancy-lactation cycle, and 2) these 2 proteins were present in milk fat globules (MFG). Expression and localization in MEC were investigated in bovine MG tissues at the end of lactation, during the dry period (DP), and early lactation using immunohistochemical and immunofluorescence approaches. The presence of ABCA1 and ABCG1 in MFG isolated from fresh milk was determined by immunofluorescence. The ABCA1 protein expression in MEC, expressed as arbitrary units, was higher during the end of lactation (12.2 ± 0.24) and the DP (12.5 ± 0.22) as compared with during early lactation (10.2 ± 0.65). In contrast, no significant change in ABCG1 expression existed between the stages. Throughout the cycle, ABCA1 and ABCG1 were detected in the apical (41.9 ± 24.8 and 49.0 ± 4.96% of cows, respectively), basal (56.2 ± 28.1 and 54.6 ± 7.78% of cows, respectively), or entire cytoplasm (56.8 ± 13.4 and 61.6 ± 14.4% of cows, respectively) of MEC, or showed combined localization. Unlike ABCG1, ABCA1 was absent at the apical aspect of MEC during early lactation. Immunolabeling experiments revealed the presence of ABCA1 and ABCG1 in MFG membranes. Findings suggest a differential, functional stage-dependent role of ABCA1 and ABCG1 in cholesterol homeostasis of the MG epithelium. The presence of ABCA1 and ABCG1 in MFG membranes suggests that these proteins are involved in cholesterol exchange between MEC and alveolar milk.     

Transfer of dietary zinc and fat to milk--evaluation of milk fat quality, milk fat precursors, and mastitis indicators

The present study demonstrated that the zinc concentration in bovine milk and blood plasma is significantly affected by the intake of saturated fat supplements. Sixteen Holstein cows were used in a 4 × 4 Latin square design with 4 periods of 12 d, and 4 dietary treatments were conducted. A total mixed ration based on corn silage, grass-clover silages, and pelleted sugar beet pulp was used on all treatments. A high de novo milk fat diet was formulated by adding rapeseed meal and molasses in the total mixed ration [39 mg of Zn/kg of dry matter (DM)], and a low de novo diet by adding saturated fat, fat-rich rapeseed cake, and corn (34 mg of Zn/kg of DM). Dietary Zn levels were increased by addition of ZnO to 83 and 80 mg of Zn/kg of DM. Treatments did not affect daily DM intake, or yield of energy-corrected milk, milk fat, or milk protein. The high de novo diet significantly increased milk fat percentage and milk content of fatty acids with chain length from C6 to C16, and decreased content of C18 and C18:1. Treatments did not influence milk free fatty acids at 4°C at 0 or 28 h after milking. The average diameter of milk fat globules was significantly greater in milk from cows offered low de novo diets. Furthermore, the low de novo diet significantly increased the concentration of nonesterified fatty acids and d-β-hydroxybutyrate in blood plasma, the latter was also increased in milk. Treatments did not affect the enzyme activity of lactate dehydrogenase and N-acetyl-β-d-glucosaminidase in milk or the activity of isocitrate dehydrogenase and malate dehydrogenase in blood plasma. The low de novo diet significantly increased plasma Zn and milk Zn content, whereas dietary Zn level did not in itself influence these parameters. This indicates that the transfer of fat from diet to milk might facilitate transfer of Zn from diet to milk.     

Elevated concentrate-to-forage ratio in dairy cow rations is associated with a shift in the diameter of milk fat globules and remodeling of their membranes

We examined the effects of concentrate-to-forage ratio in dairy cow rations on milk-fat composition, with a specific focus on the structure of milk fat globules (MFG). Twenty-four Holstein cows, 153 d in milk, were assigned to 2 dietary treatments in a crossover design study. Treatments were (1) high-concentrate (65%), low-forage (35%; HCLF) diet and (2) low-concentrate (35%), high-forage (65%; LCHF) diet. The mean diameter of the MFG; plasma concentrations of insulin, glucose, and nonesterified fatty acids (FA); and the composition and concentrations of milk FA and polar lipids were determined. Concentrations of insulin were 56% higher, and those of nonesterified FA 46% lower, in the HCLF than in the LCHF diet. The milk yield was 8.5 kg/d higher and yields of fat, protein, and lactose were 180, 350, and 403 g/d higher, respectively, in the HCLF versus LCHF diet. Milk FA composition differed between treatments, with 1.5 and 1.0 percentage units higher saturated and polyunsaturated FA concentrations, respectively, in the HCLF versus LCHF diet. Mean MFG diameter tended to be smaller (0.2 μm) in the HCLF diet than in the LCHF diet, associated with increased daily phospholipids yield (34%), lower phosphatidylserine and higher phosphatidylcholine concentrations. In conclusion, the decreased milk and fat yields in the LCHF diet were associated with remodeling of the MFG membrane and with the secretion of larger MFG. Membrane remodeling of the mammary epithelium membranes seems to play a role in regulating MFG size.     

Buffalo vs. cow milk fat globules: Size distribution,zeta-potential,compositions in total fatty acids and in polar lipids from the milk fat globule membrane

Although buffalo milk is the second most produced milk in the world, and of primary nutritional importance in various parts of the world, few studies have focused on the physicochemical properties of buffalo milk fat globules. This study is a comparative analysis of buffalo and cow milk fat globules. The larger size of buffalo fat globules, 5 vs. 3.5 μm, was related to the higher amount of fat in the buffalo milks: 73.4 ± 9.9 vs. 41.3 ± 3.7 g/kg for cow milk. Buffalo milks contained significantly lower amount of polar lipids expressed per gram of lipids (0.26% vs. 0.36%), but significantly higher amount of polar lipids per litre of milk (+26%). Buffalo and cow milk fat globule membranes contain the same classes of polar lipids; phosphatidylethanolamine, sphingomyelin (SM) and phosphatidylcholine (PC) being the main constituents. A significant higher percentage of PC and lower percentage of SM were found for buffalo milks. The fatty acid analysis revealed that saturated fatty acids, mainly palmitic acid, trans fatty acids, linolenic acid (ω3) and conjugated linolenic acid were higher in buffalo milk than in cow milk. Such results will contribute to the improvement of the quality of buffalo milk-based dairy products.     

Variation in fat globule size in bovine milk and its prediction using mid-infrared spectroscopy

The objectives of this study were to investigate the sources of variation in milk fat globule (MFG) size in bovine milk and its prediction using mid-infrared (MIR) spectroscopy. Mean MFG size was measured in 2,076 milk samples from 399 Ayrshire, Brown Swiss, Holstein, and Jersey cows, and expressed as volume moment mean (D[4,3]) and surface moment mean (D[3,2]). The mid-infrared spectra of the samples and milk performance data were also recorded during routine milk recording and testing. The effects of breed, herd nested within breed, days in milk, season, milking period, age at calving, parity, and individual animal on the variation observed in MFG size were investigated. Breed, herd nested within breed, days in milk, season, and milking period significantly affected mean MFG size. Milk fat globule size was the largest at the beginning of lactation and subsequently decreased. Milk samples with the smallest MFG on average came from Holstein cows, and those with the largest were from Jersey and Brown Swiss cows. Partial least squares regression was used to predict MFG size from MIR spectra of samples with a calibration data set containing 2,034 and 2,032 samples for D[4,3] and D[3,2], respectively. Coefficients of determination of cross validation for D[4,3] and D[3,2] prediction models were 0.51 and 0.54, respectively. The associated ratio of performance deviation values were 1.43 and 1.48 for D[4,3] and D[3,2], respectively. With these models, individual mean MFG size could not be accurately predicted, but results may be sufficient to screen samples for having either small or large MFG on average. Significant but low correlations of D[4,3] and D[3,2] with milk fat yield were estimated (0.16 and 0.21, respectively). Significant and moderate Pearson correlation coefficients for fat percent with D[4,3] and D[3,2] were assessed (0.34 and 0.36, respectively). This correlation was greater between milk fat percentage and predicted MFG size than with measured MFG size with coefficients of 0.47 and 0.49 for D[4,3] and D[3,2], respectively. The MIR prediction equations are potentially overusing the correlation between fat and MFG size and exploiting the strong relationship between the MIR spectra and total milk fat. However, the predictions of MFG size are able to determine variation in mean globule size beyond what would be achieved just by looking at the correlation with fat production.     

Effect of nonthermal technologies on the native size distribution of fat globules in bovine cheese-making milk

Milk-fat globule membranes are susceptible to damage by mechanical and thermal processes. This damage is translated into alterations of milk fat structure and functionality of cheese-making milk. The objective of this work was to evaluate the effect of pulsed electrical fields (PEF), high hydrostatic pressure (HHP), and conventional thermal treatments on fat globule size distribution and ζ-potential. Milk was processed by HHP at 400 and 500 MPa for 0–20 min, and with PEF at 36 kV/cm and 42 kV/cm up to 64 pulses. The ζ-potential of HHP and PEF treated milk were − 15.47 mV and − 14.63 mV respectively. HHP treatments induced fat globules flocculation, increasing their mass moment mean diameter. Although PEF processing did not modify the true mean diameter of MFG, it induced small globules to clump together, causing an apparent increment in the population of larger milk-fat globules.

Industrial relevance

The market for traditional raw dairy products has increased in recent times in several regions of the world due to their unique flavor and texture attributes. However, the potential negative implications of consuming raw products limit the growth of this market segment. Manufacture of raw-like cheese from thermally pasteurized milk is not feasible, among other things, because of milk fat globule membrane damage caused by elevated temperatures. Nonthermal food preservation technologies offer the potential to produce milk technically suitable for the industrial manufacture of microbiologically safe raw-like dairy products.     

乳脂消化吸收及其功能研究进展

乳脂肪球是由甘油三酯为核心的小球组成,周围有三膜结构,即乳脂肪球膜。乳脂肪球膜含有复杂的脂质和蛋白质,具有营养、免疫、神经和消化功能。然而,这些功能和胃肠消化之间存在的联系及其影响乳脂胃肠消化的因素未得到充分的研究。文章综述了乳脂成分及其结构、乳脂功能特性、乳脂胃肠消化概况及其影响乳脂消化吸收的因素,旨在为婴儿配方奶粉的消化研究提供参考。     

Changes in the surface protein of the fat globules during ultra-high pressure homogenisation and conventional treatments of milk

Disruption of fat globules upon homogenisation provokes a reduction of their size and a concomitant increase in their specific surface area. In order to overcome this phenomenon, the milk fat globule membrane (MFGM) adsorbs non-native MFGM proteins. The aim of the present study was to examine the effects of UHPH conditions (temperature and pressure) on the milk fat globule and the surface proteins by comparison with conventional treatments applied in the dairy industry. Transmission electron microscopy and SDS-PAGE revealed major differences. In UHPH-treated milk, casein micelles were found to be adsorbed on the MFGM in a lesser extent than in conventional homogenisation–pasteurisation. However, greater adsorption of directly bonded casein molecules, released by UHPH through the partial disruption of casein micelles, was observed especially at high UHPH pressures. Adsorption of whey proteins on the MFGM of conventionally homogenised–pasteurised milk was mainly through intermolecular disulfide bonds with MFGM material, whereas in UHPH-treated milk, disulfide bonding with both indirectly and directly adsorbed caseins was also involved.     

Heritabilities of measured and mid-infrared predicted milk fat globule size,milk fat and protein percentages,and their genetic correlations

The objective of this study was to estimate the heritability of milk fat globule (MFG) size and mid-infrared (MIR) predicted MFG size in Holstein cattle. The genetic correlations between measured and predicted MFG size with milk fat and protein percentage were also investigated. Average MFG size was measured in 1,583 milk samples taken from 254 Holstein cows from 29 herds across Canada. Size was expressed as volume moment mean (D[4,3]) and surface moment mean (D[3,2]). Analyzed milk samples also had average MFG size predicted from their MIR spectral records. Fat and protein percentages were obtained for all test-day milk samples in the cow's lactation. Univariate and bivariate repeatability animal models were used to estimate heritability and genetic correlations. Moderate heritabilities of 0.364 and 0.466 were found for D[4,3] and D[3,2], respectively, and a strong genetic correlation was found between the 2 traits (0.98). The heritabilities for the MIR-predicted MFG size were lower than those estimated for the measured MFG size at 0.300 for predicted D[4,3] and 0.239 for predicted D[3,2]. The genetic correlation between measured and predicted D[4,3] was 0.685; the correlation was slightly higher between measured and predicted D[3,2] at 0.764, likely due to the better prediction accuracy of D[3,2]. Milk fat percentage had moderate genetic correlations with both D[4,3] and D[3,2] (0.538 and 0.681, respectively). The genetic correlation between predicted MFG size and fat percentage was much stronger (greater than 0.97 for both predicted D[4,3] and D[3,2]). The stronger correlation suggests a limitation for the use of the predicted values of MFG size as indicator traits for true average MFG size in milk in selection programs. Larger samples sizes are required to provide better evidence of the estimated genetic parameters. A genetic component appears to exist for the average MFG size in bovine milk, and the variation could be exploited in selection programs.