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。     

Size distribution of fat globules in goat milk

Milk from French-Alpine goats and Holstein cows was obtained from a bulk tank immediately prior to analyses. Fat globule size was determined by laser particle size analysis. Individual globules of fat in goat milk ranged from 0.73 to 8.58 microm in diameter. The average diameter of particles based on volume to surface area ratio (dvs) was 2.76 microm and was less than the mean (dvs) of 3.51 microm for bovine milk, in which fat globules ranged from 0.92 to 15.75 microm in diameter. The specific surface area of particles in caprine milk was 21,778 cm2/ml, whereas the specific surface area of particles in bovine milk was 17,117 cm2/ml. Ninety percent of the total particles found in goat milk were less than 5.21 microm in diameter, whereas 90% of the total particles in bovine milk were less than 6.42 microm based on the volume frequency distribution. Dissociation of casein micelles by urea in goat whole and skim milk caused larger dvs values due to the effect of fat particles and reduced the specific surface area in both milks because the total number of detectable particles in both whole and skim milk was reduced.     

Impact of cream washing on fat globules and milk fat globule membrane proteins

Milk proteins, contained within the aqueous phase surrounding fat globules, should be removed before analysis of the composition of the native milk fat globule membrane (MFGM). The effect of the conditions applied during washing of cream on MFGM integrity has not been fully studied, and factors potentially effecting a modification of MFGM structure have not been systematically assessed so far. In this study, a cream separator was used to investigate the impact of cream washing on milk fat globule stability and the corresponding loss of MFGM proteins. Flow velocity, fat content, and type of washing solution were varied. Particle size measurements and protein analyses were carried out after each washing step to determine fat globule coalescence, removal of skim milk proteins, and efficiency of MFGM isolation. Significant differences in fat globule stability and protein amount in the MFGM isolates were measured using different washing conditions.     

Influence of enzymatic cross-linking on milk fat globules and emulsifying properties of milk proteins

The enzyme transglutaminase (TGase) can modify dairy protein functionality through cross-linking of proteins. This study examined the effects of TGase treatment on milk fat globules and the emulsifying properties of milk proteins. The extent of TGase-induced cross-linking of caseins increased with incubation time, with no differences between whole and skim milk. Extensive clustering of fat globules in extensively cross-linked raw whole milk occurred on homogenisation at 400 or 800 bar. Considerably less clustering of fat globules was observed when recombined milk (90 g fat L^–1) was prepared from TGase-treated skim milk and homogenised at 400 or 800 bar. TGase treatment did not affect fat globule size in cream, but prevented coalescence of fat globules therein, possibly through cross-linking of milk fat globule membrane components. TGase-induced cross-linking of milk proteins affected their emulsifying properties and may increase the stability of natural milk fat globules against coalescence.     

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.     

Microstructure of fat globules in whole milk after thermosonication treatment

The structure of fat globules in whole milk was studied after heat and thermosonication treatments to observe what happens during these processes at the microscopic level using scanning electron microscopy. Raw whole milk was thermosonicated in an ultrasonic processor-Hielscher UP400S (400 W, 24 kHz, 120 microm amplitude), using a 22-mm probe at 63 degrees C for 30 min. Heat treatment involved heating the milk at 63 degrees C for 30 min. Color and fat content were measured to correlate the images with analytical measurements. The results showed that the surface of the fat globule was completely roughened after thermosonication. Ultrasound waves were responsible for disintegrating the milk fat globule membrane (MFGM) by releasing the triacylglycerols. Furthermore, the overall structure of milk after sonication showed smaller fat globules (smaller than 1 microm) and a granular surface. This was due to the interaction between the disrupted MFGM and some casein micelles. Minor changes in the aspect of the globules between thermal and raw milks were detected. Color measurements showed higher L* values for sonicated samples. Sonicated milk was whiter (92.37 +/- 0.20) and generally showed a better degree of luminosity and homogenization compared to thermal treated milk (88.25 +/- 0.67) and raw milk (87.82 +/- 0.18). Fat content analysis yielded a higher value after sonication (4.24%) compared to untreated raw milk (4.04%) because fat extraction is more efficient after sonication. The advantages of thermosonicated milk are that it can be pasteurized and homogenized in just 1 step, it can be produced with important cost savings, and it has better characteristics, making thermosonication a potential processing method for milk and most other dairy products.     

Methods to assess the propensity of milk fat globules toward lipolysis and the ability of skim milk to inhibit lipolysis

Methods to quantitate factors in milk relevant to cold-induced lipolysis are described. Skim milk was incubated with milk fat globules isolated from a pool of normal milk and a fixed amount of purified lipoprotein lipase. The release of fatty acids in 24 h at 4 degrees C was determined. Most skim milk samples inhibited lipolysis, but the effect varied greatly. Skim milk from milk prone to spontaneous lipolysis was less inhibitory than skim milk from normal milk. In general, both the casein and the serum fractions of skim milk inhibited lipolysis. However, variation was greater in the effects of individual samples of milk serum. In a few extreme cases, with samples from milk with spontaneous lipolysis, the serum fraction actually stimulated lipolysis. Globules were isolated and then incubated with skim milk from normal milk and a fixed amount of purified lipoprotein lipase. This gave a measure of accessibility to lipolysis of milk fat globules in normal skim milk. There was a considerable variation in propensity toward lipolysis between milk fat globules from individual milk samples. Milk showing different levels of lipolysis obtained from five cows revealed that skim milk inhibition of lipolysis and the propensity of milk fat globules toward lipolysis were characteristic for each cow.     

Effect of the fat globule membrane on in vitro digestion of milk fat globules with pancreatic lipase

The rate and extent of in vitro lipid digestion in raw and recombined milk were investigated by determining the release of fatty acids in simulated intestinal fluid containing pancreatic lipase. Changes in the globule size, surface charge and microstructure of fat globules during digestion were examined. In the absence of bile extract, the rate of lipid digestion was slower in raw milk than in recombined milk, suggesting that the composition of the milk fat globule membrane influences the rate of lipid hydrolysis in milk. Flocculation of fat globules occurred in the early stages of digestion; the globules then coalesced to form large particles, from within which triacylglycerols were removed. However, in the presence of bile extract, the changes in the size and microstructure of the fat globules during digestion were different. Bile extract may therefore affect physicochemical interactions of fat globules and hence alter the lipolysis during the digestion.     

Behaviour of homogenized fat globules during the spray drying of whole milk

The changes in milk fat globule size and fat globule surface proteins of both low-preheated and high-preheated concentrated milks, which were homogenized at low or high pressure prior to spray drying using a disc atomization drier, were examined. The average fat globule size (d₃₂) of the spray-dried milk powders was smaller than that of the corresponding concentrates, but a small proportion of very large globules (4–80 μm) was also formed during spray drying. As a consequence, total surface protein (mg protein g⁻¹ fat) increased due to the adsorption of casein micelles at the fat globule surface during spray drying. Confocal micrographs of the powders showed some apparent spreading of the fat on the surface of the powder particles, particularly when the concentrates were homogenized at low pressure. These results indicate disruption of the milk fat globules during spray drying, which consequently causes changes in the fat globule surface protein layer.     

Size-based fractionation of native milk fat globules by two-stage centrifugal separation

A two-stage centrifugal separation method, at various separation temperatures and feed rates, was employed to fractionate milk and cream on the basis of fat globule size. It involved a modified and a conventional centrifugal separation in first and second stages, respectively. In the first stage, two streams of milk: one rich in larger fat globules and another rich in smaller fat globules, were obtained by fractionation in a modified cream separator. In the second stage, the two streams from the first stage were each further fractionated in a conventional cream separator. Depending on the temperature and feed rate of the first stage, this double separation method was able to create streams with mean fat globule size (D [4, 3]) as small as 1.35 μm and as large as 4.28 μm without affecting the droplet integrity. The developed method has potential for size-based fractionation of native fat globules in industrial scale.Industrial relevanceThe developed method has potential for size based fractionation of native fat globules in industrial scale.     

Effect of the size and interface composition of milk fat globules on their in vitro digestion by the human pancreatic lipase: Native versus homogenized milk fat globules

Although the bioavailability of dietary lipids is of primary importance in human nutrition and health, the mechanisms involved in lipid digestion are not fully understood and are of growing interest. The objective of this study was to determine the effect of the size of milk fat globules and of the composition of their interface on the activity of the human pancreatic lipase (PL). Native milk fat globules of various sizes covered by their biological membrane (MFGM) and homogenized fat globules of various sizes covered by milk proteins were prepared from whole milk and underwent lipolysis by the human PL with colipase and bile salts. A lag phase preceding the hydrolysis of milk TAG occurred with all native milk fat globules samples but not with homogenized milk samples. The kinetic parameters of human PL were determined by measuring the enzyme activity either after the lag phase for native milk fat globules samples or immediately after the addition of the enzyme for homogenized milk samples. The catalytic efficiency of human PL is 4.6-fold higher on small (1.8 μm) than large (6.7 μm) native milk fat globules, related to a 3.6-fold larger available surface. Despite the 25-fold larger available surface, milk TAG from homogenized milk are only 2-fold better hydrolyzed compared to native milk fat globules, as a possible result of a less favourable interface covered by milk proteins. The potential mechanisms involved in native vs. homogenized milk fat globules digestion by the human PL are discussed. Our study highlights the crucial role of the MFGM in the efficient digestion of milk fat globules and brings new insight for the design of dairy products and infant formulas.     

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.     

Structural changes of bovine milk fat globules during in vitro digestion

An in vitro digestion model that simulated gastric and intestinal fasting conditions was used to monitor the physical, chemical, and structural changes of fat globules from raw bovine milk. During in vitro gastric digestion, the fat globules were stable under low-acidic conditions. Some peptides and β-lactoglobulin were resistant to proteolysis by pepsin. Phospholipids, proteins, and peptides stabilized the globules in the stomach model. During in vitro intestinal digestion, most of the β-lactoglobulin and residual peptides were hydrolyzed by trypsin and chymotrypsin, and the lipolytic products, released from the hydrolysis of the triglyceride core of the globules, led to destabilization and coalescence of the globules. By accumulating at the surface of the fat globules, the lipolytic products formed a lamellar phase and their solubilization by bile salts resulted in the formation of disk-shaped micelles. This study brings new interesting insights on the digestion of bovine milk.     

Hydrolysis of bovine milk fat globules by lipoprotein lipase: inhibition by proteins extracted from milk fat globule membrane

Extraction of membrane proteins from milk fat globules by GuHCl or by MgCl2 made the lipids more accessible to lipolysis by added lipoprotein lipase. The increase in lipolysis paralleled the loss of membrane proteins and was continuous up to 2.5 M GuHCl, which was the highest concentration used. About twice as much protein was extracted with 2.5 M GuHCl as with buffer only. The amount of protein lost was about 50% of total milk fat globule protein. Lipolysis of milk fat globules was inhibited by addition of the extracted protein. The extracted proteins also reduced lipolysis when added to whole milk. More protein was needed to inhibit lipolysis of milk fat globules treated with GuHCl compared with globules treated with buffer only. The inhibition by a given amount of protein decreased if more milk fat globules were used. Protein extracted with MgCl2 had similar effects as those extracted with GuHCl. The major components extracted with MgCl2 migrated in the 40 to 50-kdalton region on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. By gel filtration chromatography, two protein fractions were obtained, which inhibited lipolysis more efficiently than the total extract. As has previously been found for inhibition of lipolysis by skim milk, the amount of extracted protein needed to inhibit lipolysis varied between preparations of milk fat globules. Milk with propensity to cold-induced ("spontaneous") lipolysis was normalized by addition of extracted proteins.     

Influence of feed composition on stability of fat globules during pumping of raw milk

Three groups of Holstein cows were fed concentrates with different fatty acid compositions; one high in saturated lipids, another high in unsaturated lipids and the last one simulating high de novo synthesis. The feedings resulted in milk with fat contents of 5.0%, 3.7% and 4.0%, respectively. The milk fat globules (MFGs) were significantly larger in the milk with the highest fat content. All three types of milk were pumped for 450 s at various shear rates and temperatures. Afterwards, measurement of particle size distribution showed that the highest coalescence of MFGs in the milk occurred with the largest fat globules. Moreover, the fat globules were more unstable at a pumping temperature of 31°C compared with lower temperatures. Likewise, an increase was found in free fatty acids for milk with the largest MFGs, indicating that milk with a high fat content is more unstable when exposed to mechanical stress. Xanthine oxidase activity in raw milk was not affected by pumping. The results are discussed in relation to automatic milking systems.     

Effect of physical properties of milk fat globules on Brucella ring test sensitivity.

Variations in Brucella milk ring test reactions have been attributed to size and disparity in size of fat globules, fat content of milk, inhibitory factors, and cream-rising capacity. Physical properties of milk fat globules were studied in individual milk samples that had diverse sensitivities for detection of Brucella agglutinins. Size or disparity of globule size could not be correlated with the milk ring test sensitivity. Inhibitory and enhancing creaming factors could be transferred from the cream to the skim milk and reacted with other cream to show changes in sensitivity. The proportion of clustered milk fat globules was related directly to the quantity of agglutinins detected by the test. Sensitivity of the test was attributed to a fat globule agglutinin that caused clustering and to an inhibitory factor.     

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.     

Oxidation of aldehydes by xanthine oxidase located on the surface of emulsions and washed milk fat globules

The enzymatic activity of xanthine oxidase (XO) located within the native and recombined milk fat globule membrane (MFGM) on emulsion surfaces was investigated after successive washing with deionised water (DW) or simulated milk ultrafiltrate (SMUF). DW was more effective than SMUF at removing non-MFMG proteins from the globule surface. Surface XO activity on a protein basis increased after three washes due to loss of MFGM proteins to the serum phase. Recombined emulsions had higher surface enzymatic activity than washed native fat globule samples. The oxidation of acetaldehyde, 3-methylbutanal, 2-methylpropanol and 2-methylbutanal to the corresponding acids by XO on the globule surface was analysed. A higher proportion of aldehydes were depleted at higher XO activity when the initial aldehyde concentration was constant, and for higher initial concentrations of aldehydes. The recovery of aldehydes and acids was above 79%, indicating that the depleted aldehydes were mostly converted to the corresponding acids.     

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.