Microfiltration of buttermilk and washed cream buttermilk for concentration of milk fat globule membrane components

Buttermilk, the by-product from butter manufacture, has gained much attention lately because of the application potential of its milk fat globule membrane (MFGM) components as health ingredients. Microfiltration (MF) has been studied for buttermilk fractionation because of its ability to separate particles from dissolved solutes. However, the presence in this by-product of skim milk solids, especially casein micelles, restricts concentration of MFGM. The use of cream washed with skim milk ultrafiltrate to produce buttermilk with lower casein content was studied as well as fractionation of this buttermilk by MF. Results have shown that washing the cream prior to churning yields buttermilk with 74% less protein than normal cream buttermilk. Analysis of the protein profile of washed cream buttermilk revealed that caseins and whey proteins were the main classes of proteins removed. The MF of washed cream buttermilk resulted in permeation fluxes 2-fold higher than with normal cream buttermilk. The second separation of the cream induced high losses of phospholipids in the skim phase. However, retention of remaining phospholipids in washed cream buttermilk by the MF membrane was higher resulting in a phospholipids concentration factor 66% higher than that of normal cream buttermilk. The results presented in this study highlight the impact of casein micelles on the separation of MFGM components as well as their effect on permeation flux during MF.     

Effect of cream treatment on phospholipids and protein recovery in butter-making process

A simple approach is proposed to recover native protein and phospholipid fractions from butter‐making process using equipments available in dairy‐processing plant. A washing treatment was used to remove protein from the cream and increase the phospholipids purity in buttermilk. Cream from a first separation was diluted with milk ultrafitration permeate and separated a second time. A 10X dilution factor reduced protein concentration in the cream from 1.6 ± 0.2 to 0.52 ± 0.03%. As a result, the phospholipids to protein ratio in buttermilk increased from 53 ± 10 to 172 ± 7 mg g^?1. In butter‐making process, an important portion of total phospholipids (～26%) is retained in butter. Butter serum made from washed cream could then be used to produce phospholipid concentrates with phospholipids to protein ratio of 473 ± 3 mg g^?1. Interestingly, the extracts from butter serum are characterised by a higher proportion of sphingomyelin compared with those from buttermilk.     

Effect of buttermilk made from creams with different heat treatment histories on properties of rennet gels and model cheeses

Although many studies have reported negative effects on cheese properties resulting from the use of buttermilk in cheese milk, the cause of these effects has not been determined. In this study, buttermilk was manufactured from raw cream and pasteurized cream, as well as from a cream derived from pasteurized whole milk. Skim milks with the same heat treatments were also manufactured to be used as controls. Compositional analysis of the buttermilks revealed a pH 4.6-insoluble protein content approximately 10% lower than that of the skim milk counterparts. Milk fat globule membrane (MFGM) proteins remained soluble at pH 4.6 in raw cream buttermilk; however, when heat was applied to cream or whole milk before butter making, MFGM proteins precipitated with the caseins. Rennet gel characterization showed that MFGM material in the buttermilks decreased the firmness and increased the set-to-cut time of rennet gels, but this effect was amplified when pasteurized cream buttermilk was added to cheese milk. The microstructure of gels was studied, and it was observed that gel appearance was very different when pasteurized cream buttermilk was used, as opposed to raw cream buttermilk. Model cheeses manufactured with buttermilks tended to have a higher moisture content than cheeses made with skim milks, explaining the higher yields obtained with buttermilk. Superior retention of MFGM particles was observed in model cheeses made from pasteurized cream buttermilk compared with raw cream buttermilk. The results from this study show that pasteurization of cream and of whole milk modifies the surface of MFGM particles, and this may explain why buttermilk has poor coagulation properties and therefore yields rennet gels with texture defects.     

Isolation of milk fat globule membrane (MFGM) material by coagulation and diafiltration of buttermilk

Due to the functional potential of milk fat globule membrane (MFGM), its isolation from buttermilk is receiving increasing attention by the food industry. However, extraction of MFGM proteins from buttermilk is challenging because of the high levels of serum proteins. In this study, a two-step approach was applied to obtain a MFGM isolate. First, native casein micelles from buttermilk were removed by rennet-induced coagulation. Next, the buttermilk whey obtained was filtered to remove the residual whey proteins. Purified MFGM isolate was collected after six diafiltration steps. The yield of MFGM proteins in the isolates was determined using sodium dodecylsulphate-polyacrylamide gel electrophoresis. In the MFGM isolates, 70% of peripheral membrane proteins were obtained, which is much more effective in comparison with recent isolation methods like cream washing or filtration of buttermilk. Further investigations on casein coagulation conditions of buttermilk may reduce losses, especially of integral MFGM proteins during renneting.     

Use of ultrafiltration and supercritical fluid extraction to obtain a whey buttermilk powder enriched in milk fat globule membrane phospholipids

Whey buttermilk, a by-product from whey cream processing to butter, is rich in milk fat globule membrane (MFGM) constituents, which have technological and potential health properties. The objective of this work was to produce a dairy ingredient enriched in MFGM material, especially phospholipids, from whey buttermilk. Whey buttermilk was concentrated by ultrafiltration (10×) and subsequently diafiltered (5×) (10 kDa molecular mass cutoff membrane) at 25 °C and the final retentate was spray-dried. The whey buttermilk powder was submitted to supercritical extraction (350 bar, 50 °C) using carbon dioxide. The membrane filtration removed most of the lactose and ash from the whey buttermilk, and the supercritical extraction extracted exclusively non-polar lipids. The final powder contained 73% protein and 21% lipids, of which 61% were phospholipids. This ingredient, a phospholipids-rich dairy powder, could be used as an emulsifier in different food systems.     

Production of a novel ingredient from buttermilk

The presence of material derived from the milk fat globule membrane (MFGM) makes buttermilk (the byproduct of butter making) distinct from any other dairy product. Membrane filtration of commercial buttermilk was carried out to obtain isolates rich in MFGM material. The separation of MFGM from the skim milk proteins present in commercial buttermilk was carried out by the addition of sodium citrate followed by microfiltration through a membrane of 0.1-microm nominal pore size. The sodium citrate caused the dissociation of casein micelles and allowed permeation of a large proportion of the skim-milk derived proteins through the membrane. This process successfully concentrated MFGM material in the retentate, and demonstrated that membrane filtration can be employed to produce MFGM fractions from commercial buttermilk. The utilization of MFGM isolates from buttermilk is of increasing importance in light of recent studies suggesting the role of phospholipids in many health-related functions: buttermilk is an untapped resource of these functional components.     

Effect of processing on the composition and microstructure of buttermilk and its milk fat globule membranes

The effect of cream pasteurization on the composition and microstructure of buttermilk after pasteurization, evaporation and spray-drying was studied. The composition of milk fat globule membrane (MFGM) isolated from buttermilk samples was also characterized. Pasteurization of cream resulted in higher lipid recovery in the buttermilk. Spray-drying of buttermilk had a significant effect on phospholipid content and composition. After spray-drying, the phospholipid content decreased by 38.2% and 40.6%, respectively in buttermilk from raw or pasteurized cream when compared with initial buttermilks. Pasteurization of cream resulted in the highest increase in whey protein recovery in MFGM isolates compared with all other processing steps applied on buttermilk. A reduction in phospholipid content was also observed in MFGM isolates following spray-drying. Transmission electron microscopy of the microstructure of buttermilks revealed extremely heterogeneous microstructures but failed to reveal any effect of the treatments.     

Compositional and functional properties of buttermilk: a comparison between sweet, sour, and whey buttermilk

Buttermilk is a dairy ingredient widely used in the food industry because of its emulsifying capacity and its positive impact on flavor. Commercial buttermilk is sweet buttermilk, a by-product from churning sweet cream into butter. However, other sources of buttermilk exist, including cultured and whey buttermilk obtained from churning of cultured cream and whey cream, respectively. The compositional and functional properties (protein solubility, viscosity, emulsifying and foaming properties) of sweet, sour, and whey buttermilk were determined at different pH levels and compared with those of skim milk and whey. Composition of sweet and cultured buttermilk was similar to skim milk, and composition of whey buttermilk was similar to whey, with the exception of fat content, which was higher in buttermilk than in skim milk or whey (6 to 20% vs. 0.3 to 0.4%). Functional properties of whey buttermilk were independent of pH, whereas sweet and cultured buttermilk exhibited lower protein solubility and emulsifying properties as well as a higher viscosity at low pH (pH ≤ 5). Sweet, sour, and whey buttermilks showed higher emulsifying properties and lower foaming capacity than milk and whey because of the presence of milk fat globule membrane components. Furthermore, among the various buttermilks, whey buttermilk was the one showing the highest emulsifying properties and the lowest foaming capacity. This could be due to a higher ratio of phospholipids to protein in whey buttermilk compared with cultured or sweet buttermilk. Whey buttermilk appears to be a promising and unique ingredient in the formulation of low pH foods.     

Whipping properties of recombined,additive-free creams

There is increasing industrial interest in the use of the milkfat globule membrane as a food ingredient. The objective of this research was to determine whether the aerosol whipping performance of cream separated into butter and buttermilk, and then recombined, would perform in a manner similar to untreated cream. Churning of cream tempered to different solid fat contents was used to separate butter from buttermilk, which were then recombined at the same ratios as the initial extraction yield, or with 25% extra buttermilk. Differences in milkfat globule size distributions among the recombined creams were apparent; however, their whipping behavior and overrun were similar. Importantly, all recombined creams did not yield properties similar to the original cream, indicating that the unique native milkfat globule membrane structure plays a role in cream performance well beyond its simple presence.     

Casein precipitation by acid and rennet coagulation of buttermilk: Impact of pH and temperature on the isolation of milk fat globule membrane proteins

Milk fat globule membrane (MFGM) proteins make up only 2–3% of the total protein content in buttermilk. Prior isolation of buttermilk MFGM material is therefore required for further investigation of its functional properties and application in food systems. Rennet induced coagulation has been applied to remove caseins, but 30–70% of MFGM proteins are entrapped in the casein curd under classical cheese making conditions. Therefore, factors influencing the rennet as well as acid induced coagulation of buttermilk with regard to a reduction of MFGM protein losses into the casein curd, i.e., pH, temperature and pre-heat treatment of buttermilk, were examined. Complete casein removal from buttermilk was reached using both coagulation methods, whereas retention of MFGM proteins in the serum phase was higher during renneting. Rennet induced coagulation at higher pH and lower temperatures resulted in weaker gel structures, which increased the yield of MFGM proteins in the buttermilk whey.     

β-Casein-phospholipid monolayers as model systems to understand lipid-protein interactions in the milk fat globule membrane

Phospholipid-protein monolayer films were studied as model systems to mimic the structure of the native bovine milk fat globule membrane (MFGM) and to understand lipid-protein interactions at the surface of the globule. Phospholipids extracted from bovine raw milk, raw cream, processed milk and buttermilk powder were spread onto the air-water interface of a Langmuir trough, β-casein was then added to the sub-phase, and Langmuir-Blodgett films were studied by epifluorescence microscopy and atomic force microscopy. In all films, β-casein was responsible for clustering of the sphingomyelin- and cholesterol-rich microdomains into larger platforms. This suggests that the same phenomenon may happen at the surface of the milk fat globule, where specific MFGM proteins may cause aggregation of microdomains.     

Phospholipid enrichment in sweet and whey cream buttermilk powders using supercritical fluid extraction

Milk fat globule membrane contains many complex lipids implicated in an assortment of biological processes. Microfiltration coupled with supercritical fluid extraction (SFE) has been shown to provide a method of concentrating these nutritionally valuable lipids into a novel ingredient. In the dairy industry there are several by-products that are rich in phospholipids (PL) such as buttermilk, whey, and whey cream. However, PL are present at low concentrations. To enrich PL in buttermilk powders, regular buttermilk and whey buttermilk (by-product of whey cream after making butter) were microfiltered and then treated with SFE after drying. The total fat, namely nonpolar lipids, in the powders was reduced by 38 to 55%, and phospholipids were concentrated by a factor of 5-fold. Characterization of the PL demonstrated specific molecular fatty amide combinations on the sphingosine (18:1) backbone of sphingomyelin with the greatest proportion being saturated; the most common were 16:0, 20:0, 21:0, 22:0, 23:0, and 24:0. Two unsaturated fatty amide chains, 23:1 and 24:1, were shown to be elevated in a whey cream buttermilk sample compared with the others. However, most unsaturated species were not as abundant.     

Microfiltration of butter serum upon casein micelle destabilization

The gross composition of butter serum, the aqueous phase of butter, is comparable to that of buttermilk, except that it has a higher content of material derived from the milk fat globule membrane (MFGM). As such, butter serum is a good source for further purification of MFGM material. The purified fraction could be of interest for its emulsifying and nutritional properties. The effect of sodium citrate and ethanol on the dissociation of butter serum casein micelles, and their effect on casein retention upon tangential microfiltration were investigated. Optimal conditions of casein micelle dissociation were assessed by using an experimental design (response surface full central composite orthogonal design) with temperature and ethanol or sodium citrate concentration as design variables and the Hunter L* value as response variable. For both dissociating agents, a highly significant reduced quadratic model was fit to the data. Microfiltration tests were performed on pure butter serum, and on butter serum in the presence of sodium citrate, under optimal dissociation conditions (50°C, 80 mM). A cellulose acetate membrane with a pore size of 0.15 μm was used. From the filtration curves and fouling coefficients it was clear that the addition of sodium citrate improved the permeation flux, and minimized fouling. All fractions were analyzed for dry matter, protein, lactose, lipid, and polar lipid contents. The protein fraction was further characterized by sodium dodecyl sulfate-PAGE. It was shown that sodium citrate greatly enhanced casein transmission through the membrane, but at the expense of substantial losses of polar lipids.     

Effect of heating whey proteins in the presence of milk fat globule membrane extract or phospholipids from buttermilk

The objective of this study was to determine the contribution of phospholipids from buttermilk as a nucleus in the heat-induced aggregation of whey proteins. Solutions of whey proteins (5%, w/v) were adjusted to pH 4.6 or 6.8 and then heated at 65 or 80 °C for 25 min with or without 1% (w/v) of milk fat globule membrane (MFGM) extract or phospholipid powder. The aggregation mechanisms were characterised using analysis with Ellman's reagent, one-dimensional gel electrophoresis, thin-layer chromatography, and three-dimensional confocal laser-scanning microscopy. Three-dimensional images showed protein/phospholipid interactions in the presence of MFGM extract or phospholipids, and thin-layer chromatography plates showed no trace of free phospholipids after 20 min at pH 4.6. Overall, the results demonstrate that phospholipids from buttermilk were involved in the formation of protein aggregates through the MFGM fragments at a low temperature, whereas phospholipids could interact directly with the proteins at a higher temperature (80 °C).     

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.     

Effect of washing conditions on the recovery of milk fat globule membrane proteins during the isolation of milk fat globule membrane from milk

During the isolation of milk fat globule membrane (MFGM) from milk, washing is considered the most critical stage in which loss of MFGM components occurs. In this study, using a cream separator, the influence of washing on the recovery of MFGM proteins was investigated. The residue of non-MFGM proteins in the MFGM material obtained after washing was quantitatively determined using densitometric analysis of one-dimensional sodium dodecyl sulfate-PAGE after silver staining of the gel. Using deionized water as the washing solution did not increase the loss of MFGM proteins compared with other common salt solutions in terms of recovery of MFGM proteins and contamination with non-MFGM proteins. The increase in wash temperature from 38 to 46°C did not show a significant decrease in yield of MFGM proteins because of variation between the experimental replicates. Coalescence of fat globules occurs during isolation. To increase MFGM purity while maintaining a high MFGM protein recovery, using larger volumes of wash solution is more advisable rather than increasing the number of washings from 2 to 3.     

Concentration of polar MFGM lipids from buttermilk by microfiltration and supercritical fluid extraction

Buttermilk contains the milk fat globule membrane (MFGM), a material that possesses many complex lipids that function as nutritionally valuable molecules. Milk-derived sphingolipids and phospholipids affect numerous cell functions, including regulating growth and development, molecular transport systems, stress responses, cross membrane trafficking, and absorption processes. We developed a two-step method to produce buttermilk derivative ingredients containing increased concentrations of the polar MFGM lipids by microfiltration and supercritical fluid extraction (SFE). These processes offer environmentally benign alternatives to conventional lipid fractionation methods that rely on toxic solvents. Firstly, using a ceramic tubular membrane with 0.8-micron pore size, we evaluated the cross flow microfiltration system that maximally concentrated the polar MFGM lipids using a 2n factorial design; the experimental factors were buttermilk source (fresh, or reconstituted from powder) and temperature (50 degrees C, and 4 degrees C). Secondly, a SFE process using supercritical carbon dioxide removed exclusively nonpolar lipid material from the microfiltered buttermilk product. Lipid analysis showed that after SFE, the product contained a significantly reduced concentration of nonpolar lipids, and a significantly increased concentration of polar lipids derived from the MFGM. Particle size analysis revealed an impact of SFE on the product structure. The efficiency of the SFE system using the microfiltration-processed powder was compared much more favorably to using buttermilk powder.     

Filtration of milk fat globule membrane fragments from acid buttermilk cheese whey

The proteins and polar lipids present in milk fat globule membrane (MFGM) fragments are gaining attention for their technological and nutritional properties. These MFGM fragments are preferentially enriched in side streams of the dairy industry, like butter serum, buttermilk, and whey. The objective of this study was to recover MFGM fragments from whey by tangential filtration techniques. Acid buttermilk cheese whey was chosen as a source for purification by tangential membrane filtration because it is relatively rich in MFGM-fragments and because casein micelles are absent. Polyethersulfone and cellulose acetate membranes of different pore sizes were evaluated on polar lipid and MFGM-protein retention upon filtration at 40°C. All fractions were analyzed for dry matter, ash, lipids, proteins, reducing sugars, polar lipid content by HPLC, and for the presence of MFGM proteins by sodium dodecyl sulfate-PAGE. A fouling coefficient was calculated. It was found that a thermocalcic aggregation whey pretreatment was very effective in the clarification of the whey, but resulted in low permeate fluxes and high retention of ash and whey proteins. By means of an experimental design, the influence of pH and temperature on the fouling and the retention of polar lipids (and thus MFGM fragments), proteins, and total lipids upon microfiltration with 0.15 μM cellulose acetate membrane was investigated. All models were highly significant, and no outliers were observed. By increasing the pH from 4.6 to 7.5, polar lipid retention at 50°C increased from 64 to 98%, whereas fouling of the filtration membrane was minimized. A 3-step diafiltration of acid whey under these conditions resulted in a polar lipid concentration of 6.79 g/100 g of dry matter. As such, this study shows that tangential filtration techniques are suited for the purification of MFGM fragments.     

Thermocalcic aggregation of milk fat globule membrane fragments from acid buttermilk cheese whey

Fragments originating from the milk fat globule membrane (MFGM), which is rich in polar lipids and membrane-specific proteins, are gaining interest for their functional and nutritional properties. Acid buttermilk cheese whey was used as a source for MFGM purification, because its MFGM content is more than 5 times higher than that of standard rennet whey. Because polar lipids are the main constituent of the MFGM and only occur in membranous structures, the polar lipid content was taken as a parameter for the total MFGM fragment content. The process of thermocalcic aggregation was evaluated on its recovery of MFGM fragments in the pellet. This method, originally intended for whey clarification and defatting, is a combination of calcium addition, a pH increase, and a thermal treatment. The influence of pH (6.5 to 8), temperature (40 to 70°C), and calcium concentration (0.1 to 0.24 g/100 g) on the pellet mass and dry matter (DM) content and on recovery of protein and polar lipids (and thus indirectly on MFGM fragments) was investigated by means of a response surface Box-Behnken orthogonal design. Reduced quadratic models were fit to the experimental data and were found to be highly significant. No outliers were observed. The recovery of MFGM fragments was found to be highly dependent on the pH, and less dependent on temperature and calcium addition. Next to MFGM proteins, whey proteins were also found to be involved in the formation of aggregates. Optimal conditions were found at 55°C, pH 7.7, and 0.205 g of calcium/L of whey. Under these conditions, 91.0% of the whey polar lipids were recovered in a firm and compact pellet of only 7.86% of the original whey mass, with a polar lipid concentration of 8.34% on pellet DM. Washing with water and centrifugation of the pellet was successful because after one washing step, virtually all sugars were removed, whereas 75.9% of the whey polar lipids could still be recovered. As such, the polar lipid content of the washed pellet increased to 10.70% on a DM basis. However, a second washing step resulted in serious losses of MFGM material.