Comparative study of the molecular species of chloropropanediol diesters and triacylglycerols in milk fat

In an effort to establish the origin of the fatty acid esters of 3-chloropropanediol, which recently have been isolated in small amounts from goat milk, we compared the molecular species composition of the chlorohydrin diesters and of goat milk triacylglycerols. The chloropropanediol diesters were found to be composed of molecular species containing C₁₀−C₁₈ fatty acids and corresponded closely in carbon number to those calculated for the long chain sn-1,2-diacyl-glycerol moieties of goat milk triacylglycerols. The molecular species of goat milk total triacylglycerols contained C₄−C₁₈ fatty acids. It is suggested that triacylglycerols and chloropropanediol diesters are derived from the same pool of long chain fatty acids. A molecular distillate of bovine milk fat did not contain chloropropanediol diesters, while the available samples of human milk fat were shown to contain alkyldiacylglycerols as the major components of a neutral lipid fraction corresponding in polarity to the chloropropanediol diesters.     

TAG composition of ewe's milk fat. Detection of foreign fats

The TAG composition of 45 samples of ewe's milk, collected throughout the year from five Spanish breeds, was analyzed according to their carbon number by using short capillary column GC. The TAG content had a bimodal distribution with maxima at C₃₈ (12.8%) and C₅₂ (8.4%). The TAG composition did not vary significantly with respect to the time of year of sampling but was affected by the breed. Multiple regression equations based on TAG content are proposed to detect foreign fats in ewe's milk fat. Analysis of known mixtures of lard, palm oil, and cow's milk fat with ewe's milk fat have experimentally confirmed the accuracy of the equations.     

Prediction of the goat milk fatty acids by near infrared reflectance spectroscopy

Near infrared reflectance spectroscopy (NIRS) was used to predict the goat milk fatty acid (FA) profile. The ability of cow milk broad‐based calibration equations to predict the goat milk FA profile was assessed. Three hundred twenty‐eight samples in the calibration set and 108 in the validation set were analyzed. We showed that the bias and unexplained error were significant for most of the FA despite an adequate standardized Mahalanobis distance (index to establish the boundaries of a population of samples) which allowed us to test the ability of bovine models to predict goat milk FA profiles. To better predict the goat milk FA composition, a specific goat model was investigated. The cross‐validation coefficient of determination (R²CV; proportion of variance explained by the model in cross‐validation) and residual predictive deviation (RPD; index which allows to standardize the standard error of prediction (SEP)) were >0.90 and 3, for saturated, monounsaturated, and unsaturated FA, total trans FA, isomer cis9trans11 of CLA, cis9‐, trans10‐, and trans11‐C18:1, respectively. Monitoring of the equation performance of milk FA included the calculation of the bias and unexplained error. Practical applications: In this work, we evaluated the ability of NIRS to predict FA composition of goat milk and tested the ability of using broad‐based cow milk calibration equations to monitor an independent set of goat milk samples. We tested the hypothesis that the calibration equations obtained for FA from cow milk could be applied on goat milk. As the spectra of goat milk are similar to those of cow milk when they are measured by the standardized Mahalanobis distance (index to establish the boundaries of a population of samples), we can accept the hypothesis. It will be possible to use the existing calibration equations for predicting FA profile on spectra of milk of a different specie. However, the rejection of the hypothesis would put in doubt the use of Mahalanobis distance as the unique index for testing the performance of calibration equations on different milk populations for predicting FA.     

Triglyceride composition of bovine milk fat with elevated levels of linoleic acid

The effect of increasing the linoleic acid (18∶2) content of milk fat on the composition and structure of the triglycerides (TG) was investigated. Protected sunflower seed supplement was added to the diet of a cow grazing on pasture, and the structure and composition of the milk fat compared with the milk fat from its monozygous twin which had been fed a control diet. The relative proportions of TG fractions of high, medium, and low molecular weight in the milk fat with elevated levels of 18∶2 (15.5% 18∶2) were 43.0, 19.5, and 37.5 moles %, respectively, compared with 36.1, 19.7, and 44.2 moles %, respectively, in the milk fat from the cow fed the control diet. Separation of these three TG fractions of each milk fat into TG classes with different levels of unsaturation showed that the milk fat with elevated levels of 18∶2 contained higher proportions of diene, triene, and tetraene TG and correspondingly lower proportions of saturated and, to a lesser extent, monoene TG. The saturated and monoene TG from the two milk fats had similar fatty acid compositions. However, the diene TG of the 18∶2-rich milk fat included high proportions of the combination of 18∶2 with two saturated fatty acids (FA) which are minor constituents of normal milk fats. Likewise, the triene TG reflected the presence of 18∶2 in combination with 18∶1 and a saturated FA.     

Specific distribution of fatty acids in the milk fat triglycerides of goat and sheep

The triglycerides of the fat globules of sheep and goat milk were isolated and separated into short and long chain lengths by silicic acid column chromatography. The short chain lengths comprised major triglycerides with 34–44 acyl carbon atoms and accounted for nearly 50% of the total milk fat. The long chain lengths contained major triglycerides with 40–54 acyl carbons. Stereospecific analyses of the short chain triglyceride fraction showed that of the 20–23 moles per cent of C₄−C₈ fatty acids present, at least 95% were specifically attached to the glycerol molecule in the position corresponding to carbon 3 ofsn-glycerol. The distribution of the other fatty acids (C₁₀ or greater) did not show such marked specificity for either the 1 or the 2 position. Although individual triglycerides were not identified, the specific placement of the fatty acids could best the accounted for by assuming a common pool of long chain 1,2-diglycerides which served as precursors of the bulk of both short and long chain triglycerides during milk fat synthesis. Presented in part at the AOCS Meeting, New York, October 1968.     

HPLC‐APCI analysis of triacylglycerols in milk fat from different sources

Triacylglycerols (TAG) from milk‐fat from different sources (cow, goat and human milks) were characterised using non‐aqueous reversed phase high‐performance liquid chromatography–atmospheric‐pressure chemical ionisation, coupled to MS/MS (RP HPLC‐APCI MS/MS). The fatty‐acid content of all samples was also established by methyl transesterification and GC‐MS analysis. Optimisation of the HPLC gradient, combined with APCI tandem MS, enables TAGs complex mixtures to be analysed without prior separation. More than 160 different glycerides were identified; between 50 and 70 compounds were identified in the chromatograms of each sample. This method also enabled the principal TAG regioisomers to be recognized. The study focused on the investigation of the structure of TAGs containing very‐long‐chain PUFA, namely all cis‐ 4,7,10,13,16,19‐ DHA (DHA, C_{22:6, n‐3}) and all cis‐5,8,11,14,17‐eicosapentaenoic acid (EPA, C_{20:5, n‐3}), both in human and in n‐3‐enriched cow's milks. Ten TAGs containing DHA were identified in human milk and only one in milk from cows fed an n‐3 enriched diet.     

Cholesterol esters of milk and mammary tissue

The fatty acid composition and metabolic activity of cholesterol esters in milk and mammary tissue (cow, sow and goat) were investigated. Cholesterol esters of freshly secreted milks incubated at 40 C for 2 hr showed no change in fatty acid composition and no incorporation of 1-¹⁴C-palmitate. The fatty acid composition of cholesterol esters in the milk of all three species exhibited elevated levels of a unique group of fatty acids when compared to other milk ester lipid classes. This group was comprised of monounsaturated acids and acids with odd numbers of carbons. Tissue cholesterol esters contained lower levels of these acids. Evidence from experiments in which an odd carbon acid (C₁₅) was infused into the lactating mammary gland indicated that the group of unique acids is preferentially retained in the cholesterol ester fraction which is secreted with milk. These infusion experiments also provided evidence that cholesterol esters accumulate in developing milk fat globules in a manner paralleling triglyceride accumulation, and that acyl moieties of cholesterol esters may be desaturated in the form of the intact ester. Our results are compatible with the hypothesis that acyl moieties of cholesterol esters in lactating mammary tissue are turning over rapidly. Paper No. 3496 in the journal series of the Pennsylvania Agricultural Experiment Station.     

Effect of outlet drying temperature and milk fat content on the physicochemical characteristics of spray-dried camel milk powder

Abstract

A composite face-centered experimental design was used to investigate the influence of spray drying conditions on the physicochemical characteristics of camel and cow milk powders. Response surface methodology (RSM) was deployed to appraise the effects of these processing parameters (the outlet drying temperature and the milk fat content) on water activity (a_w), glass transition temperature (T_g), bulk density, and free fat quantity. According to RSM analysis, it was noticed that the a_w and the T_g were primarily influenced by the outlet drying temperature instead of by milk fat content. Our results highlighted the negative effects of milk fat content and of the outlet drying temperature on the bulk density as well as on the free fat quantity of camel milk powder. Likewise, our findings underlined the negative effect of the outlet drying temperature on the bulk density of cow milk powder. However, the increase of fat content has led to the overexposure of fat at the free surface of the cow milk powder. Our results suggested a marked similarity of the overall thermodynamic behavior of both milks, during drying. Nevertheless, some differences were highlighted regarding the structuring of the particles of camel milk powder.     

Production and characterization of a swiss cheese-like product from modified vegetable oils

Seven types of Swiss cheese-like products were made by recombining skim milk with various fat sources: higholeic sunflower oil (HOSO), milk fat, randomized milk fat, HOSO with commerical short-chain fatty acids (C₄−C₁₀) (SCFA) interesterified at 100 and 120% of the levels in the milk fat, HOSO with interesterified milk fat SCFA, and HOSO with dissolved free SCFA. Sensory, chemical, and physical analyses were conducted to evaluate the flavor and texture of the cheeses. All cheeses made from HOSO with interesterified SCFA were not significantly different from milk fat controls in typical Swiss flavor and volatile flavor. HOSO with interesterified SCFA scored significantly higher in these flavors than unmodified HOSO. Swiss flavor was positively correlated with sweetness (0.805), volatile flavor (0.737), caramelized flavor (0.703), non-fat solids (0.663), and SCFA (0.639), and negatively correlated with fat (−0.645) and salt (−0.482) content. A linear regression model was established for typical Swiss flavor that included fat, salt, titratable acidity, and medium- and long-chain fatty acids as variables (R ²=0.91). Instrumental texture profile analysis indicated no differences among the treatments in texture atributes except cohesiveness. The production of a good-flavored Swiss cheeselike product from HOSO with interesterified SCFA appears to be commercially feasible.     

Asymmetric distribution of decanoate in triacylglycerol synthesized in vitro by mammary glands of lactating mice

Slices, prepared from the mammary glands of lactating mice, were incubated with either [1-¹⁴C]acetate, [U-¹⁴C]glucose, or [1-¹⁴C]decanoate. From all 3 substrates, radioactivity in the synthesized lipids was found mainly in triacylglycerols (TG). When acetate or glucose served as substrate, decanoate (C₁₀) accounted for 24% of the fatty acids in TG. Hydrolysis of the TG by pancreatic lipase yielded [¹⁴C] fatty acids which had relatively more C₁₀ (38%) than did either of the other hydrolysis products mono- or diacylglycerol (14–17%). However, when TG produced by slices from C₁₀ were hydrolyzed, the acid was found to be esterified equally at the C-1, C-2 and C-3 of glycerol. Thus, when fatty acids are synthesized de novo and are converted to TG by gland slices, C₁₀ is predominantly located in the C-3 position, a finding in accord with the situation in milk TG, although such preferential incorporation does not occur when the free acid is presented to the tissue slices.     

Content and distribution oftrans-18:1 acids in ruminant milk and meat fats. Their importance in european diets and their effect on human milk

Thetrans-18:1 acid content and distribution in fats from ewe and goat milk, beef meat and tallow were determined by a combination of capillary gas-liquid chromatography and argentation thin-layer chromatography of fatty acid isopropyl esters. Thetrans isomers account for 4.5 ± 1.1% of total fatty acids in ewe milk fat (seven samples) and 2.7±0.9% in goat milk fat (eight samples). In both species, as in cow, the main isomer is vaccenic (trans-11 18:1) acid. The distribution profile oftrans-18:1 acids is similar among the three species. The contribution of ewe and goat milk fat to the daily intake oftrans-18:1 acids was estimated for people from southern countries of the European Economic Community (EEC): France, Italy, Greece, Spain, and Portugal. It is practically negligible for most of these countries, but in Greece, ewe and goat milk fat contributeca. 45% of the daily consumption oftrans-18:1 acids from all dairy products (0.63 g/person/day for a total of 1.34 g/person/day). Thetrans-18:1 acid contents of beef meat fat (ten retail cuts, lean part) and tallow (two samples) are 2.0 ± 0.9% and 4.6%, respectively, of total fatty acids (animals slaughtered in winter). Here too, the main isomer is vaccenic acid. Othertrans isomers have a distribution pattern similar to that of milk fat. Beef meat fat contributes less than one-tenth of milk fat to thetrans-18:1 acid consumed. The daily per capita intake oftrans-18:1 acids from ruminant fats is 1.3–1.8 g for people from most countries of the EEC, Spain and Portugal being exceptions (ca. 0.8 g/person/day). In France, the respective contributions of ruminant fats and margarines to the daily consumption oftrans-18:1 acids are 1.7 and 1.1 g/person/day (60 and 40% of total, respectively). These proportions, based on consumption data, were confirmed by the analysis of fat from milk of French women (ten subjects). The mean content oftrans-18:1 acids in human milk is 2.0 ± 0.6%, with vaccenic acid being the major isomer. Based on the relative levels of thetrans-16 18:1 isomer, we could confirm that milk fat is responsible for the major part of the daily intake oftrans-18:1 acids by French people. The daily individual intake oftrans-18:1 isomers from both ruminant fats and margarines for the twelve EEC countries varies from 1.5 g in Spain to 5.8 g in Denmark, showing a well-marked gradient from the southwest to the northeast of the EEC.     

Triglyceride composition of ewe,cow, and goat milk fat

The separation and identification of the components in milk fat, which are mainly triglycerides, is a challenge due to its complex composition. A reverse-phase high-performance liquid chromatography (HPLC) method with gradient elution and light-scattering detection is described in this paper for the triglyceride analysis in ewes’ milk fat. Triglyceride identification was carried out by combining HPLC, gas-liquid chromatography (GLC), and the calculated equivalent carbon numbers of several triglyceride standards. Quantitation of partially resolved peaks in the HPLC chromatogram was accomplished by applying a peak deconvolution program. Forty-four fatty acids were identified by GLC analysis, but only 19 were used for the following prediction of triglyceride molecular species; 181 triglycerides were identified, some of which were grouped at the same peak and needed application of the deconvolution program. Consequently, coefficients of variation were close to or lower than 5%. Moreover, the triglyceride composition of ewe, cow, and goat milk fat were compared by using these methods. These results show that ewe milk fat is richer in short- and medium-chain triglycerides, and cow milk fat is richer in long-chain and unsaturated triglycerides.     

Milk fat structure of a patient with type 1 hyperlipidemia

Structural analyses were performed on milk fat samples obtained 3–10 days postpartum from a lactating patient with primary Type 1 hyperlipidemia. The milk triacylglycerols contained 3–7% C₁₀, 14–21% C₁₂, 20–30% C₁₄, 22–26% C₁₆ and 20–30% C₁₈ (largely oleic) acids. Gas liquid chromatographic (GLC) analyses of the X-1,3- and X-1,2-diacylglycerols on polar siloxane columns showed a markedly non-random association of acyl chains. Stereospecific analyses indicated that the short chain length fatty acids were confined essentially to the sn-3-position of the triacylglycerol molecule. Furthermore, these acids were largely absent from the phosphatidylcholines and the endogenous sn-1,2-diacylglycerols of the milk fat. It is concluded that the short chain fatty acids are incorporated into the milk triacylglycerols during the final stage of biosynthesis via the phosphatidic acid pathway, and that the overall fatty acid distribution is consistent with the 1-random 2-random 3-random hypothesis.     

Ceramide structure of sphingomyelin from human milk fat globule membrane

Sphingomyelin was purified from human milk fat globule membrane and submitted to phospholipase C to yield ceramide. The structure of this ceramide was investigated by gas liquid chromatographic analyses of its components, fatty acids and sphingoid bases. The structure of the native ceramide was confirmed by direct-inlet mass spectrometry. It was shown to contain a major base C₁₈-sphingosine associated with a high proportion (60%) of C₂₀, C₂₂, C₂₄ and C_24∶1 nonhydroxylated fatty acids. As these very long-chain fatty acids might be of nutritive importance, the concentration of sphingomyelin in human milk and its distribution in cream and skim milk were established.     

Phase equilibrium and crystallization behavior of mixed lipid systems

As complex lipid systems, the phase and crystallization behavior of mixtures of a high-melting milk fat fraction with a low-melting milk fat fraction or canola oil was studied. A turbidity technique was developed to estimate solubility and metastability conditions of these lipid mixtures. Both solubility and metastability of the high-melting milk fat fraction in liquid lipids increased exponentially with temperature. At a given equilibration temperature, liquid phases and solid fractions with nearly identical melting profiles and TAG compositions were obtained regardless of the original concentration of the lipid mixture. The maximum melting temperature (MMT), as measured by DSC, of the liquid phase increased dramatically in the equilibrium temperature range of 27.5–35.0°C but did not change at temperatures below and above this range (down to 25.0°C and up to 40°C in this study). The content of long-chain TAG (C₄₆−C₅₂) increased and short-chain TAG (C₃₆−C₄₀) decreased in the liquid phases as the equilibrium temperature increased. A plot of the TAG group ratio (i.e, long-short-chain TAG) vs. equilibrium temperature was generated to illustrate the phase behavior of the complex lipid system and to represent a solubility curve, from which the supersaturation level for crystallization kinetics was determined. Higher supersaturation and lower temperature resulted in higher nucleation and crystallization rates. Compared to the system with a low-melting milk fat fraction, mixtures of the high-melting milk fat fraction with canola oil had higher nucleation and crystallization rates due to the lower solubility found for this system.     

Occurrence of Aflatoxin M1 in Dairy Products in Southern Italy

A screening survey of the presence of aflatoxin M₁ (AFM₁) was carried out on 265 samples of cheese made from cow, buffalo, goat, sheep, sheep-goat milk collected in the Apulia region (Southern Italy). Selected samples included unripened, medium and long-term ripened cheeses. AFM₁ was found in 16.6% of the analyzed samples. The highest positive incidence was for medium and long-term ripened cheeses, especially those made from sheep-goat milk, while buffalo cheeses tested consistently negative. Our results show that the level of contamination by AFM₁ in dairy products from Apulia Region are lower than in other Italian and European regions. Moreover, it is important to underline that a common European norm concerning the AFM₁ threshold limits for dairy products is still lacking.     

Structure of bovine milk fat triglycerides: II. Long chain lengths

The long chain triglycerides of bovine milk fat were isolated by thin layer chromatography, and their chemical structure determined by combined thin layer and gas liquid chromatography, and a stereospecific analysis of a molecular distillate of butteroil of comparable composition. The milk fat fraction (39% of total) contained C₈–C₂₀ fatty acids which were distributed among the glycerides of 40–56 acyl carbon atoms in a manner not unlike that found for the same acids in the short chain triglycerides. Although individual triglycerides were not identified, the specific distribution of the fatty acids could best be accounted for by assuming a common pool of long chain 1,2-diglyceride precursors from which the bulk of both short and long chain triglycerides are synthesized by a stereospecific introduction of C₄–C₁₈ fatty acids in position 3 of sn-glycerol. This hypothesis is compatible with the results of stereospecific analyses of the short and long chain fractions and of the total butteroil. It is supported by the nonrandom distributions demonstrated for the molecular weights of the milk fat triglycerides of different degrees of saturation. Presented in part at the AOCS meeting, Philadelphia, October, 1966.     

Postparturition changes in the triacylglycerols of cow colostrum

The changes in the triacylglycerol (TAG) composition of colostrum fat of three cows were studied. In addition to the determination of fatty acid composition by gas chromatography, the distribution of TAG according to the acyl carbon number (ACN) and molecular weight was analyzed utilizing both supercritical fluid chromatography (SFC) and ammonia negative-ion chemical ionization mass spectrometry (MS). Colostrum TAG contained substantially less stearic and oleic acids and more myristic and palmitic acids than the normal Finnish milk fat. The major trends in the changes of fatty acids and TAG were similar for each cow, although clear differences between individuals were found. During the first week of parturition, the proportions of short-chain fatty acids (C₄–C₁₀) typically increased as well as those of stearic and oleic acids, whereas the relative amounts of C₁₂–C₁₆ acids decreased, especially those of myristic and palmitic acids. Distinct changes occurred also in TAG distributions: the proportions of molecules with ACN 38–40 increased and those with ACN 44–48 decreased. Although there were distinct differences between individuals shortly after delivery, both the fatty acid compositions and TAG distributions of the milk samples of the cows started to resemble each other after one week. The theoretical profiles of colostrum TAG calculated based on the fatty acid compositions differed clearly from the ACN distributions analyzed by SFC and MS. Thus, the analysis of TAG is essential, because the changes in molecular species composition of colostrum TAG cannot be estimated according to the fatty acid analysis alone.     

Supercritical Carbon Dioxide Fractionation of Anhydrous Milk Fat

Supercritical carbon dioxide was used to fractionate anhydrous milk fat. Six fractions were produced at 40, 50 and 60 °C using pressure values of 10, 20, 25, 30, 33 and 36 MPa. The fractions were analyzed for fatty acids, thermal behavior, iodine and color values. Composition and yield of fatty acid methyl esters were evaluated at different fractionation conditions in relation to the original milk fat values. Short chain fatty acids (C₄–C₈), medium chain fatty acids (C₁₀–C₁₄) and total saturated fatty acids were decreased from fraction obtained in the order of 10–36 MPa, while long chain fatty acids (C₁₆–C_18:2) and total unsaturated fatty acids were increased. Fractions obtained in the raffinate stage of the fractionation exhibited higher melting behavior that obtained at the low CO₂ pressures. The higher iodine value of raffinate fraction indicated that fraction was richer in oleic acid. Fractions produced at low pressures had lower melting behavior than those obtained at high pressures. Yellowness Index and b* values increased in raffinate fraction due to concentration of carotenoids.