Untersuchungen über die Kristallstruktur und den molekularen Aufbau der Fettphase von Butter

Studies on the Crystal Structure and the Molecular Conformation of the Fat Phase of Butter Comparative examinations of butter samples are carried out at different temperatures of preparation ranging from 15 to 35° C by means of electron microscope pictures won by freezefracturing technique and of X-ray diffraction patterns. While an increase of temperature from 15 to 25–30° C causes a liquefaction of the crystalline aggregates, contained in the interior of the fat globules, the main part of the outer globule shell remains unchanged. However, in case of a further increase of temperature to 35° C practically all fat globules and crystals in the interglobular fat phase of butter melt. Nevertheless, many water droplets, existing isolated with their characteristic covering of small and thin crystal platelets, still remain in existence. It is concluded that in the region of the outer globule shell as well as in the region of the crystal platelets of the water droplets the triclinic β modification dominates. In contrast to this fact the predominating part of the β′ modification is to be found in the deeper melting crystal layers of the interior of the globules. This is true for the interglobular fat phase, too, in which the part of unsaturated fatty acids is especially high. Further on the arrangements of molecules in the β and β3′ regions of butter are discussed, whereby the numerous lattice defects are particularly mentioned.     

The fatty acid composition of small and large naturally occurring milk fat globules

Native milk fat globules of various mean diameters, ranging from d₄₃ = 1.5 to 7.3 μm, were obtained using microfiltration of raw whole milk acquired in winter and spring. After total lipid extraction, fatty acid composition was characterized by methyl and butyl ester analysis using gas chromatography. The oleic and linoleic acid content of milk obtained in winter increased with fat globule size, whereas myristic and palmitic acid decreased. There was significantly more lauric, myristic and palmitoleic acid, and less stearic acid in small fat globules compared to large fat globules in milk obtained in both winter and spring. The relative content of oleic and linoleic acids were found to depend on fat globule size and season. Results are interpreted on the basis of the relative content of milk fat globule membrane depending on fat globule size, and on consequences of compositional variations on milk fat globule melting behavior.     

Milk fat globules: Fatty acid composition,size and in vivo regulation of fat liquidity

Populations of large and small milk fat globules were isolated and analyzed to determine differences in fatty acid composition. Globule samples were obtained by centrifugation from milks of a herd and of individual animals produced under both pasture and barn feeding. Triacylglycerols of total globule lipids were prepared by thin layer chromatography and analyzed for fatty acid composition by gas chromatography. Using content of the acids in large globules as 100%, small globules contained fewer short-chain acids, −5.9%, less stearic acid, −22.7%, and more oleic acids, +4.6%, mean values for five trials. These differences are consistent with alternative use of short-chain acids or oleic acid converted from stearic acid to maintain liquidity at body temperature of milk fat globules and their precursors, intracellular lipid droplets. Stearyl-CoA desaturase (EC 1.14.99.5), which maintains fluidity of cellular endoplasmic reticulum membrane, is suggested to play a key role in regulating globule fat liquidity. Possible origins of differences between individual globules in fatty acid composition of their triacylglycerols are discussed.     

Differently sized native milk fat globules separated by microfiltration: fatty acid composition of the milk fat globule membrane and triglyceride core

The aim of this study was to characterize the fatty acid composition of the core and membrane of differently sized milk fat globules separated by microfiltration, which can now be used to manufacture dairy products. Native milk fat globules of various mean diameters, ranging from d₄₃ = 2.3 µm to 8.0 µm, were obtained using microfiltration of raw whole milk. After milk fat globule washing, the milk fat globule membrane (MFGM) and the triglyceride core (TC) were separated by manual churning. After total lipid extraction from each fraction, their fatty acid composition was characterized using methyl ester analysis by gas chromatography. Regardless of season, no significant differences were observed in the fatty acid composition of the MFGM phospholipids. Conversely, significant differences were found in the fatty acid composition of TC; particularly, small fat globule TC contained more medium‐chain fatty acids and less stearic acid than large fat globule TC. These results show that the previously observed differences in total fatty acid composition among differently sized milk fat globules are due to their triglyceride composition and MFGM amount rather than to the composition of the MFGM.     

Zur fütterungsbedingten Variation des Milchfettes der Kuh unter besonderer Berücksichtigung von Regulationsmechanismen beim Fettstoffwechsel

Variations of Cow's Milk Fat as Affected by Feeding with Special Consideration of Regulation Mechanisms of Fat Metabolism Milk fat composition is highly affected by the conditions of feeding, which again may have considerable consequences for the quality of several dairy products. In order to establish possible variations in physical and chemical properties of the milk fat as depending on the nutritional status of the cow, a small herd of cows was successively subjected to different defined feeding conditions (barn feeding, pasture feeding, energy deficit). The investigations indicated that from barn feeding to pasture feeding to energy deficit the proportions of oleic acid and of long chain triglycerides increased and the proportions of short and medium chain fatty acids and triglycerides decreased. In spite of highly differing fat compositions relatively constant values for the viscosity and the average triglyceride molecular weight were obtained; also various characteristics from the melting and solidification curves were nearly identical. The solid/liquid ratios approached at higher temperatures. These results point to the existence of regulation mechanisms, in the course of which the flow and melting properties are kept within the required narrow ranges by an opposite behaviour of the proportions of short chain fatty acids and oleic acid. With pasture feeding, compared with barn feeding, all oleic acid isomers and linoleic acid increased, with energy deficit the proportions of these acids are at the low level of barn feeding. These results as well as the continuous decrease of the cholesterol content from barn to pasture to energy deficit are explained on the basis of the different pathways of fat metabolism; deficit are explained on the basis of the different pathways of fat metabolism; furthermore the proportions of trans C18: 1 acids are discussed in connection with the milk fat content. The milk fat globules are particularly large and unstable at the period of energy deficit. Consequently, quality deterioration of whipping cream (creaming, plug formation) and other dairy products (lipolysis) can be expected.     

The Effect of Phospholipids on Milkfat Crystallization Behavior

Depending on the production method, the fat component in butter can either be in a continuous phase or entrapped by milk fat globule membranes (MFGM). Phospholipids like those present in the MFGM are known to affect milk fat crystallization behavior. This study examines the effect of MFGM phospholipid concentration on butter crystal structure. Regular raw cream was first concentrated to 85% fat via centrifugation to produce a “plastic” cream, and was then blended with anhydrous milk fat (AMF) and skim milk in order to maintain the total fat-to-water ratio while altering the total amount of MFGM in the butter product. Whereas mixtures of AMF and skim milk contained large spherulites that had finer structures as the degree of supercooling was increased, the addition of globular fat (GF) broke up these structures, eventually producing smaller individual needle-like crystals. However, high levels of GF also lead to the coalescence of the aqueous phase, creating large water pockets that adversely affect sensory properties. Thus, the phospholipid concentration in the final butter product must be controlled in order to obtain optimal crystal structure and product quality.     

Design of a Microstructured System for the Homogenization of Dairy Products at High Fat Content Part II: Influence of Process Parameters

Partial homogenization using a microstructured SHM (Simultaneous Homogenizing and Mixing) valve significantly reduces aggregation of fat globules within their homogenization by feeding the continuous phase directly into the droplet disruption zone (as discussed in part I of this work). It allows homogenization of cream containing up to at least 42 vol.‐% fat, and thus, significantly reduces processing costs without loss in product quality, i.e., overall process intensification. The present article details current results on the influence of material parameters, i.e., emulsifier system and fat content, and process parameters, i.e., homogenizing pressure and the temperature of both streams, on the fat globule size distribution. In contrast to conventional technology, SHM valve technology generates decreasing droplet sizes with increasing pressure or with increasing temperature of the homogenizing stream. This novel technique breaks new ground in dairy product design.     

A method for isolation of milk fat globules

The traditional procedure for isolating milk fat globules involves repeated cycles of centrifuging to obtain globules and redispersion of them in fresh buffer to eliminate other milk components. We have evaluated a simpler, less manipulative method whereby globules are centrifuged out of the milk and through an overlying buffer layer. Human milk samples ranging from 0.1 to 35 ml were centrifuged at 1500×g for 20 min after deposition under a suitable quantity of buffer. This yielded purified globules, in less time, which could be dispersed more satisfactorily than those by the traditional procedure. Protein, phospholipid and cholesterol contents of globules by the two methods were quite similar. A lower protein content (10.4 vs 13.2 mg/g of lipid) was characteristic of globules prepared by the multiple wash method. However, large differences could not be seen in gel electrophoresis patterns of the proteins. By using plastic centrifuge tubes, tube freezing and cleavage just below the globule layer enables clean separation of globule and nonglobule phases for analysis of milk component distributions. Macro (5 to 35 ml of sample) and micro (200 μl or less) versions of the method are described. Limited trials showed that the method can be applied satisfactorily to cow's and goat's milks, but for highly pure globules a deeper buffer column than that used with human milk is required because of their much higher casein content.     

Lipid domains in the milk fat globule membrane: Specific role of sphingomyelin

Lipids are secreted in milk in the form of unique colloidal assemblies, the milk fat globules, which are surrounded by a biological membrane, the milk fat globule membrane (MFGM). Recent confocal laser scanning microscopy experiments showed heterogeneities in the composition and structure of the MFGM. For the first time, we revealed the phase separation of polar lipids in the plane of the MFGM with the lateral segregation of sphingomyelin and cholesterol in rigid liquid‐ordered (L_o) domains surrounded by the fluid matrix of glycerophospholipids in the liquid‐disordered (L_d) phase. The MFGM is a unique and highly dynamic biophysical system, the functional and nutritional properties of which need to be further studied for health benefits.     

Membrane phospholipids and sterols in microfiltered milk fat globules

Native milk fat globules of various mean diameters, ranging from d₄₃ = 2.3 µm to 8.0 µm, were obtained using microfiltration of raw whole milk. After milk fat globule washing, the milk fat globule membrane (MFGM) was separated by manual churning. After total lipid extraction and separation of polar lipids, their phospholipid (PL) and sterol composition was measured using thin‐layer chromatography, methyl ester analyses by gas chromatography, and gas chromatography coupled to mass spectrometry. The main PL species were phosphatidylethanolamine, phosphatidylcholine and sphingomyelin. The respective fatty acid composition of each PL species was measured. Many different minor bioactive sterols were detected in the MFGM, e.g. lanosterol, lathosterol, desmosterol, stigmasterol and β‐sitosterol. No significant differences in the PL and sterol profile were found between MFGM extracted from small and large milk fat globule fractions.     

Induction of Milkfat Crystallization in the Emulsified State by High Hydrostatic Pressure

The effect of high hydrostatic pressure to milk fat in the emulsified state has been studied by transmission election microscopy (TEM) and low resolution pulsed nuclear magnetic resonance (NMR) for the purpose of following pressure-induced crystallization processes. Pasteurized dairy creams (35 and 43% fat) were subjected to pressures between 100 and 500 MPa for 1 to 15 minutes at a temperature of 23°C. By applying the freeze fracture technique for TEM studies it was demonstrated that pressurization induces fat crystallization within the emulsion droplets, predominantly at the globule periphery. The ultrastructural findings were confirmed by NMR which showed that the amount of crystallized fat formed depended both on intensity and time of pressurization. In the pressurized cream samples the crystallization proceeded during further storage at 23°C whereas the fat droplets in the original cream sample remained liquid under these conditions, provided that the cream sample was held at this temperature.     

Emulsifying properties of phospholipids in the reconstitution of cream using butter oil

Phospholipids (PL) extracted from bovine milk and soybeans were tested for their emulsifying properties in reconstituted cream using butter oil. Bovine milk PL dispersed in the oil phase were found to stabilize the cream, whereas soybean PL were found to solidify the cream. Differential scanning calorimetry was carried out on the two creams, revealing that the addition of soybean PL to the butter oil retarded the onset of crystallization during cooling, which consequently led to a lower solid fat content within the fat globules. It was suggested that the difference in the crystallization behavior of butter oil due to the addition of different PL led to the different emulsion stabilities of the reconstituted creams.     

A comparative study of the lipids of globule membrane and fat core and of the milk serum of cows

Nine samples of fresh raw cow's milk were separated into fat globules and milk serum by centrifugation. After destabilization by freezing and thawing, the milk fat globules were resolved into membranes and fat cores. The lipid composition of these structures was compared to that of the surrounding milk serum. Of the total milk fat, 95–98% was in the fat cores, 0.5–1% in the globule membranes and the rest (1.5–4%) in the milk serum. The fat cores contained 88–93% triglyceride, 5.2–9.8% diglyceride, 1.5–7.3% free fatty acid and 0.2–0.4% cholesterol, but no phospholipid. The lipids of the membrane contained 21–44% phospholipid, made up of about equal proportions of phosphatidyl ethanolamine, phosphatidyl choline, and sphingomyelin. The other lipids of the membrane (56–79%) consisted of 83–88% triglyceride, 5.1–10.7% diglyceride, 1–5.1% free fatty acid and 0.4–1.9% cholesterol. The milk serum contained 30–45% phospholipid divided about equally among phosphatidyl ethanolamine, phosphatidyl choline and sphingomyelin. The rest (55–70%) of the milk serum lipids was made up of 71–83% triglycerides, 4.3–10.1% diglycerides, 8.7–15.7% free fatty acids, and 1.2–8.4% cholesterol. Corresponding phospholipid classes of milk serum and globule membranes had identical fatty acid compositions. The triglycerides and diglycerides of the globule membranes possessed increased proportions of palmitic and stearic acids in comparison to the glycerides of the fat cores. Taken in part from a PhD thesis submitted by T. C. Huang to Queen's University, Kingston, Canada in April, 1965. Presented in part at the 47th Canadian Chemical Conference and Exhibition held in Kingston, Canada, June 1–3, 1964.     

The turnover time of dietary cholesterol in the lactating rat

The secretion of dietary 4-¹⁴C-cholesterol into milk of the rat was determined as a function of post-feeding time by a single dose technique. The time interval which elapsed before maximum specific radioactivity was reached in milk (17–20 hr after maximum activity in the serum) suggests a route through the mammary gland involving transport of the cholesterol by intracellular membranes. It also suggests that the exogenous cholesterol is incorporated into the milk fat globule membranes rather than into the fat globules during their synthesis within the cell. Paper No. 3978 in the Journal Series of the Pennsylvania Agricultural Experiment Station.     

Effects of Emulsifiers on Protein-Fat Interaction in Ice Cream Mix during Ageing I: Quantitative Analyses

Ageing of ice cream mix at 0 to 10°C for a minimum of 3 up to 24 hours prior to ice cream production in the ice cream machine is generally used to improve various physical properties. During ageing the fat phase of the mix crystallizes and protein is slowly desorbed from the surface of the fat globules. In the presence of emulsifiers fat crystallization and protein desorption is distinctly enhanced resulting in increased agglomeration of fat globules during ice cream manufacture which is important for the formation and stability of the final ice cream. The effect of emulsifiers on ageing is due to their strong interfacial activity at low temperature.     

Milk – A new source for bioactive phospholipids for use in food formulations

The fat globules of milk are surrounded by a membrane, which contains a triple layer of phospholipids. Concentrated cream is thus a very good raw material for further processing in order to make a powder, enriched in phospholipids. A powder can in this way easily be produced with 20% phospholipids using only common dairy processes such as centrifugation and membrane filtration. Milk phospholipids differ in compositions from existing commercial sources such as soybean lecithins and egg lecithins particularly with respect to the content of two bioactive phospholipids namely sphingomyelin (about 24%) and phosphatidylserine (12%). Sphingomyelin is an active agent controlling the intestinal uptake of cholesterol and triacylglycerols in human nutrition. Intake of phosphatidyl serine has clinically documented effects in maintaining cognitive performance like memory and stress control. A milk phospholpids enriched product thus offers a 100% natural alternative to the well‐established, semi‐synthetic head‐group exchanged phosphatidyl serine from soy lecithin now existing on the market for functional foods formulations.     

Crystallization and fractionation of milk fat

Recent progress in understanding milk fat crystallization and fractionation is reviewed. Extent of fat solidification in butter can be altered by variations in thermal treatment of cream prior to churning. Because of its compositional complexity, milk fat rarely exhibits polymorphism. As with mixtures of closely related triglycerides, milk fat forms solid solutions. A typical milk fat begins melting below −40 C, maximum melting occurs at 15–18 C, and the highest melting fraction appears 20–37 C as a shoulder on the main peak. Dispersion of fat in emulsions increases its tolerance to supercooling, thereby altering the properties and composition of the solid phase. Most studies of milk fat fractionation have used progressive fractional crystallization, either of the melt or of solutions. Both procedures result in fractions showing larger changes in mp than in composition. The high melting glyceride fraction, ca. 5% total fat, influences crystallization out of proportion to concentration. The Alfa-Laval system, using an aqueous suspension of partially crystalline fat, produces two fractions. Typical high melting fractions have softening points ca. 3C higher than the original fat. The softening point of typical low melting fractions is lowered 10 C. Refractionation is easier with the high melting fraction. Melting thermograms of these fractions show them as resembling fractions prepared from melted fat. One of eight papers presented at the Symposium “Milk Lipids,” AOCS Fall Meeting, Ottawa, Canada, September 1972.     

Origin of the milk fat globule

Milk fat globules originate as fat droplets within the lactating mammary cell. These droplets are composed largely (>98%) of glycerides. Their constituent fatty acids are derived by lipolysis of very low density lipoproteins and chylomicrons of the blood and by the novo synthesis within the cell. Evidence of two principal routes for the synthesis of milk fat triglycerides has been presented: the so-called glycerolphosphate and monoglyceride pathways. Recent findings on these pathways are discussed. The nature of the milk triglycerides with their unique compliment and distribution of short chain fatty acids appears to depend upon a closely regulated relation between the soluble multienzyme complex that synthesizes the fatty acids and the glyceride synthetase that is bound to the endoplasmic reticulum. The resulting triglycerides appear to self-assemble into droplets from the surface of the endoplasmic reticulum. No special ultrastructures (transport particles, vesicles, etc.) have been detected in relation to this process. Milk fat droplets at the time of secretion average several microns in diameter, there being species variations. The basic secretion mechanism involves envelopment of the droplet in plasma membrane and expulsion of it from the cell. As a consequence there are at least two pools of polar lipids (cholesterol and phospholipids) associated with secreted milk fat globules, i.e., one from the plasma membrane and one entrained earlier from the endoplasmic reticulum at the time of triglyceride synthesis and accumulation. In all, the polar lipids do not make up more than 1–2% of the total lipids in milk and a substantial fraction of them has been identified recently with plasma membrane fragments occurring in the skim milk phase. Radiotracer and ultrastructural studies show that this membrane material does not result simply by shedding of surface from milk fat globules. This dispersed material and the lining around milk fat globules constitute valuable sources for the study of cell membranes. One of eight papers presented in the symposium “Milk Lipids,” AOCS Meeting, Ottawa, September 1972.     

Der Einfluß von Emulgatoren auf das Verhalten von Fett im Eismix während des Reifens

Effect of Emulsifiers on the Behaviour of Fat in Ice Cream Mix During Ageing Ageing of ice cream mix takes up most of the time in the production of ice cream. This is due to the fat crystallisation which is important for the processing of the mix in the freezer. The influence of emulsifiers is investigated with regard to their degree of saturation and their hydrophilic-lipophilic balance using simple laboratory methods such as the determination of the free extractable fat and the emulsion stability. Saturated emulsifiers do not show any remarkable changes of the extractable fat and the emulsion stability during the ageing time. Saturated emulsifiers cause an increase of the extractable fat and a deterioration of the emulsion stability. These changes are especially obvious during the first three to six hours of ageing. The definite orientation of the protein-emulsifier membrane around the fat globules is only finished after twelve to twenty-four hours of ageing. This might explain the fact that in practice three hours of ageing are considered as absolutely necessary and that a longer ageing time causes a further improve of ice cream quality.     

Role of actinidin in the hydrolysis of the cream milk proteins

Cream from milk is commonly used as the starting material of variety food products such as cream cheese, fat added cheeses, butter and buttermilk. The protein content of cream is only about 2% but strongly affects the quality of fat containing dairy products especially cheeses which are frequently characterized by an unsatisfactory texture. In this work, the suitability of the actinidin to function in a “fat-supplemented” environment was tested and the SDS–PAGE profile of cream treated with the enzyme was also analyzed. The results show that actinidin is able to degrade all the cream proteins to smaller than 29.0 kDa fragments suggesting a novel utilization as preparatory treatment in the production of milk derived products characterized by a reduced content of undesirable proteins.