Fat Triacylglycerols Research Articles

Production of Human Milk Fat Substitutes by Lipase-Catalyzed Acidolysis: Immobilization, Synthesis, Molecular Docking and Optimization Studies

Human milk fat (HMF) triacylglycerols (TAGs) mainly contain palmitic acid esterified at the sn-2 position while oleic and other unsaturated fatty acids are located at positions sn-1,3. This study aimed at the production of HMF substitutes (HMFS) by lipase-catalyzed acidolysis of tripalmitin with oleic acid, in a solvent-free medium. Burkholderia cepacia lipase (BCL) was immobilized in silica (prepared with protic or aprotic ionic liquids) by covalent binding or encapsulation and used as biocatalyst. The supports and immobilized biocatalysts were characterized by FTIR, TGA, and SEM. Molecular docking analysis showed that BCL preferentially attacks oleic acid rather than tripalmitin, due to the lower free energy of hydrophobic binding with this acid (−6.5 kcal·mol−1) than with tripalmitin (5.4 kcal·mol−1). Therefore, the tripalmitin attack by BCL and subsequent HMFS production only occurs after the binding to most of the oleic acid molecules. The highest acidolysis activity was obtained with BCL immobilized by covalent binding in prepared silica with aprotic ionic liquid. A central composite rotatable design, as a function of temperature (58–72 °C) and oleic acid/tripalmitin molar ratio (MR = 2:1–6.8:1), was performed for acidolysis optimization. Under optimized conditions (58 °C and MR = 4:1 or 60 °C and MR = 2:1), the oleic acid incorporation of 28 mol.% was achieved after 48 h.

A novel UHPLC-ESI-MS/MS method and automatic calculation software for regiospecific analysis of triacylglycerols in natural fats and oils

Regioisomeric analysis of triacylglycerols (TAGs) in natural oils and fats is a highly challenging task in analytical chemistry. Here we present a software (TAG Analyzer) for automatic calculation of regioisomeric composition of TAGs based on the mass spectral data from recently reported ultra-high performance liquid chromatography electrospray ionization tandem mass spectrometry (UHPLC−ESI−MS/MS) method for analyzing TAG regioisomers. The software enables fast and accurate processing of complex product ion spectra containing structurally informative diacylglycerol [M+NH4−RCO2H–NH3]+ and fatty acid ketene [RCO]+ fragment ions. Compared to manual processing, the developed software offers higher throughput with faster calculation as well as more accurate interpretation of chromatographically overlapping isobaric TAGs. The software determines results by constructing a synthetic spectrum to match the measured fragment ion spectrum, and by reporting the optimal concentrations of TAGs used to create the synthetic spectrum. This type of calculation is often extremely challenging for manual interpretation of the fragment ion spectra of isobaric TAGs with shared fragments, hence the need for automated data processing. The developed software was validated by analyzing a wide range of mixtures of regiopure TAG reference compounds of known composition and a commercial olive oil sample. Additionally, the method was also applied for regiospecific analysis of TAGs in human milk as an example of natural fats and oils with a highly complex TAG profile. The results indicate that the software is capable of resolving regioisomeric composition of natural TAGs even of the most complex composition. This novel calculation software combined with our existing UHPLC-ESI-MS/MS method form a highly efficient tool for regioisomeric analysis of TAGs in natural fats and oils.

Feeding hydrogenated palm fatty acids and rumen-protected protein to lactating Holstein-Friesian dairy cows modifies milk fat triacylglycerol composition and structure, and solid fat content

The aim of this study was to analyze the effect of fat and protein supplementation to dairy cattle rations on milk fat triacylglycerol (TAG) composition, fatty acid (FA) positional distribution in the TAG structure, and milk solid fat content (SFC). Fifty-six lactating Holstein-Friesian cows were blocked into 14 groups of 4 cows and randomly assigned 1 of 4 dietary treatments fed for 28 d: (1) low protein, low fat, (2) high protein, low fat, (3) low protein, high fat, and (4) high protein, high fat. The high protein and high fat diets were obtained by isoenergetically supplementing the basal ration (low protein, low fat) with rumen-protected soybean meal and rumen-protected rapeseed meal, and hydrogenated palm FA (mainly C16:0 and C18:0), respectively. Fat supplementation modified milk TAG composition more extensively compared with protein supplementation. Fat supplementation resulted in decreased concentrations of the low molecular weight TAG carbon number (CN) 26 to CN34 and medium molecular weight TAG CN40, CN44, and CN46, and increased concentrations of CN38 and the high molecular weight TAG CN50 and CN52. Increased contents of C16:0, C18:0, and C18:1cis-9 in TAG in response to fat supplementation were related to increases in the relative concentrations of C16:0 and C18:0 at the sn-2 position and C18:0 and C18:1cis-9 at the sn-1(3) positions of the TAG structure. Increased concentrations of high molecular weight TAG species CN50 and CN52 in response to fat supplementation was associated with increased milk SFC at 20, 25, and 30°C. Our study shows that important alterations in milk TAG composition and structure occur when feeding hydrogenated palm FA to lactating dairy cattle, and that these alterations result in an increased SFC of milk fat. These changes in milk SFC and TAG composition and structure may improve absorption of both fat and minerals in milk-based products for infants and may affect processing of milk fat.

Synthesis of margarine fat from sesame oil and palm stearin by chemical interesterification

The production of margarine fat is not only intended to be free from Trans Fatty Acid (TFA) but is also expected to have a higher quality from a nutritional aspect. In this research, margarine fat from sesame oil (SO) and palm stearin (PS) by a chemical interesterification was synthesized. Chemical interesterification is one of the processes used to modify the physico-chemical characteristics of oils and fats. An attempt to chemical-restructure palm stearin and sesame oil to form margarine fat which is suitable for margarine was investigated using sodium methoxide as a catalyst. The effect ratio of PS/SO in color, slip melting point, solid fat index, texture and triacylglycerols (TAGS) profile of margarine fat were studied in research. This research was conducted by three major stages; characterization of sesame oil and palm stearin, synthesis of margarine fat by physical blending and chemical interesterification, and characterization of margarine fat. This study used a Completely Randomized Design (CRD) with one factor, namely the ratio of sesame oil and palm stearin. Margarine fat produced with different variations of the raw material concentration (% w/w SO:PS = 30:70; 40: 60; 50:50, 60:40, and 70:30). Chemical interesterification caused: rearrangement of triacylglycerols, reduction of S3 and U3 and increase in S2U and U2S type TAGs content of all blend, resulting in lowering of melting point, solid fat index and increase texture. Margarine fat in the % ratio of SO:PS = 60:40, has a similar characteristic (texture, slip melting point, solid fat index, and TAGS profiles) which margarine commercial, so has the potential to be developed in the manufacture margarine industry

Effect of season, feeding, and anatomical region on the triacylglycerol profile of Iberian pig fat

The work studies the effects of season, feeding type, and anatomical region on the Iberian pig fat triacylglycerol (TAG) profiles, considered as compositional data (CoDa). The analysis consisted of applying exploratory tools in the simplex and standard multivariate techniques to data transformed into the Euclidean space (ilr coordinates). Compositional biplot showed differences in TAG containing palmitic (P) and oleic (O) acids between the 2005 and 2003/2004 seasons but not within these. PPP (clr variance, 0.139), OLL (0.095), PPS (0.075), POPo + PLP (0.074), and PSS (0.629) showed high CoDa variability among treatments. The ANOVA analysis found significant (p ≤ 0.05) effects of season, feeding type, and anatomical region on pig fat TAG profile, but only that for 2005 season was well predicted (97.5%) by discriminant analysis (DA). Overall, season was more influential on the Iberian pig fat TAG profile than montanera length and sampling region, which effects were not significant for some statistical techniques.

Effect of pasteurisation, homogenisation and freeze‐drying on bovine and buffalo milk fat triacylglycerols profile

Milk fat triacylglycerols (TAGs) analysis is considered to be a challenging task; considerable numbers of fatty acyl moieties form one of the most complicated natural fats, with potentially thousands of molecular species of TAGs. Herein, TAGs fingerprinting of raw and processed bovine and buffalo milk was carried out using ultra‐performance liquid chromatography‐mass spectrometry. Firstly, milk samples were subjected to pasteurisation, homogenisation and freeze‐drying, and the TAGs profile was then analysed. Palmitic, oleic, myristic and stearic were the predominant fatty acids detected. The results revealed that the TAG profile was not significantly influenced by the applied technological treatments. Bovine milk TAGs exhibited 28–54 total acyl carbons, with 0–5 double bonds at 80 different retention times. However, buffalo milk TAGs possessed 26–54 total acyl carbon number containing 0–4 double bonds at 84 different retention times. The findings of this study could offer valuable knowledge to the field of dairy technology.

Strategy for stereospecific characterization of natural triacylglycerols using multidimensional chromatography and mass spectrometry

Stereoisomeric determination of individual triacylglycerols (TAGs) in natural oils and fats is a challenge due to similar physicochemical properties of TAGs with different fatty acid combinations. In this study, we present a strategy to resolve the enantiomeric composition of nutritionally important TAGs in sea buckthorn (Hippophaë rhamnoides) as an example food matrix. The targeted strategy combines 1) fatty acid profiling with GC, 2) separation of TAGs with RP-HPLC, 3) stereospecific separation with chiral-phase HPLC and 4) structural characterization with MS. Three major asymmetric diacid- and triacid-TAG species were analyzed in sea buckthorn pulp oil. Off-line coupling of RP-HPLC and chiral-phase HPLC allowed separation of several TAG regioisomers and enantiomers, which could not be resolved using one-dimensional techniques. Enantiomeric ratios were determined and specific structural analysis of separated TAGs was performed using direct inlet ammonia negative ion chemical ionization method.Of the TAG 16:0/16:1/16:1 palmitic acid (C16:0) was located predominantly in a primary position and the enantiomeric ratio of TAG sn-16:1-16:1-16:0 to sn-16:0-16:1-16:1 was 70.5/29.5. Among the TAGs 16:0/16:0/18:2 and 16:0/16:0/16:1, only ca 5% had C16:0 in the sn-2 position, thus, ca 95% were symmetric sn-16:0-18:2-16:0 and sn-16:0-16:1-16:0. The enantiomeric ratio of triacid-TAGs containing C16:0 and two unsaturated fatty acids (palmitoleic C16:1, oleic C18:1 or linoleic acids C18:2) could not be resolved due to lack of commercial enantiopure reference compounds. However, it became clear that the targeted strategy presented offer unique and convenient method to study the enantiomeric structure of individual TAGs.

Computational simulation of nanostructured lipid carrier containing lipids from Cupuassu (Theobroma grandiflorum) seed fat: Design, interaction and molecular dynamic study

Drug delivery systems are constantly evolving and developing, as well as the search for promising and effective formulations for drug delivery. Computational simulation methods enable the development of complex systems, such as nanostructured lipid carriers (NLC), the understanding of interaction and dynamics between drug molecule and its transporter. In this work, aimed to simulate a NLC containing cupuassu fat triacylglycerols, carnauba wax and caprylic/capric acid triacylglycerol, stabilized with Tween 80 and Pluronic and ketoconazole enantiomer as drug was simulated. Initially, lipid mixtures were studied by Differential Scanning Calorimetry and X-ray diffraction. Subsequently, computational studies were carried out, among which Molecular Docking of ketoconazole to the lipid mixture and Molecular Dynamics of NLC system containing ketoconazole. From the results obtained it was possible to observe the main binding affinities of the drug and provide a better NLC formulation. It was also possible to propose a three-dimensional NLC model that was stable after molecular dynamics and ideal for future experimental studies.

The effect of cooling on crystallization and physico-chemical properties of puff pastry shortening made of palm oil and anhydrous milk fat blends

Commercial shortenings and margarines are mainly produced via rapid cooling with scraped surface heat exchangers (SSHE) and therefore, controlling fat crystallization in the cooling process of these systems plays an important role. In this study, four batches of puff pastry shortening were produced from palm oil blends containing 25% and 50% anhydrous milk fat (AMF) using a bench-top SSHE. The outlet temperature of SSHE was investigated at 20 °C and 12 °C relating to the difference of crystallization temperature between palm oil and milk fat. Physico-chemical properties, crystal morphology and fat stability of these shortenings during storage at 5 °C were determined by various classical technologies including differential scanning calorimetry (DSC), polarized light microscopy (PLM), rheological and texture analyzer. Our results showed that fat blends crystallized at 12 °C had a stronger crystal network after production but the discrepancy of hardness diminished after the first storage week at 5 °C. Besides, the change of crystallization temperature did not affect significantly melting behavior as well as fat polymorphism of products. Upon storage, some big aggregates were detected in shortenings as a combined result of the recrystallization of palm fat crystals and the post-crystallization of milk fat triacylglycerols (TAGs). • Fat blends crystallized at 12 °C had a stronger crystal network than at 20 °C. • The discrepancy of hardness between fat blends diminished in storage at 5 °C. • Crystallization temperature (12 °C–20 °C) did not affect polymorphism of products. • Post-crystallization of AMF promoted the formation of big aggregates in products.

Physicochemical Properties of Palm Kernel Oil and Palm Kernel Olein Blends

Palm kernel oil (PKO) and palm kernel olein (PKOo) have limited application due to rapid crystallisation properties and excessive foaming during heating, respectively. In this study, a series of PKO and PKOo blends were prepared at different concentration and analysed for slip melting point (SMP), solid fat content (SFC), triacylglycerols (TAGs) composition, thermal and morphological properties. The addition of 90% RBDPKOo caused the SMP to decrease from 27.8°C to 23.80°C. All the samples show steep SFC melting profile from 0°C to 30°C. The onset crystallisation temperature of RBDPKO was delayed with the addition of 90% PKOo from 9.73°C to 5.80°C and totally melted at lower melting temperatures. It was observed that the addition of RBDPKOo caused the crystals to be spherulitic, scattered and less aggregated with each other. The addition of RBDPKOo reduced the compositions of the main TAG of RBDPKO such as LaLaLa, LaLaM and CLaLa. Therefore, through blending the physicochemical properties of the RBDPKO and RBDPKOo could be changed. Hence, their applications in food could be widened.

Appliance of a high pressure semi-batch reactor: supercritical transesterification of soybean oil using methanol

Abstract This work presents the results of a supercritical transesterification process of soybean oil using methanol, performed in a laboratory high pressure semi-batch reactor. A full description of the equipment and a standard operating procedure are also provided. Since fossil fuels are not renewable, biofuels such biodiesel, have gained attention over the past decades. Biodiesel is the result of the transesterification of triacylglycerols in vegetable oils or animal fats and it works as an alternative fuel for compression-ignition engines. Methanol was chosen to perform the reaction, at molar ratios of 1:24 (oil: alcohol) and 0.1 (CO 2:methanol), temperature of 280 °C and pressure of 130 bar. In order to decrease the reaction time, CO2 in the supercritical state was used as cosolvent. The yield of biodiesel produced was equal to 95.19% in only ten minutes of reaction. A high yield of methyl esters (84.41%) was detected by gas chromatography-mass spectrometry. In general, 10 compounds were identified. The component in higher concentration was linoleate, followed by methyl hexadecanoate, oleic acid methyl ester and methyl stearate. The ester conversion rate means that the process with supercritical methanol can be successfully applied to produce biodiesel.

Lipid Profile, Lipase Bioactivity, and Lipophilic Antioxidant Content in High Pressure Processed Donor Human Milk.

Human milk fat plays an essential role as the source of energy and cell function regulator; therefore, the preservation of unique human milk donors’ lipid composition is of fundamental importance. To compare the effects of high pressure processing (HPP) and holder pasteurization on lipidome, human milk was processed at 62.5 °C for 30 min and at five variants of HPP from 450 MPa to 600 MPa, respectively. Lipase activity was estimated with QuantiChrom™ assay. Fatty acid composition was determined with the gas chromatographic technique, and free fatty acids content by titration with 0.1 M KOH. The positional distribution of fatty acid in triacylglycerols was performed. The oxidative induction time was obtained from the pressure differential scanning calorimetry. Carotenoids in human milk were measured by liquid chromatography. Bile salt stimulated lipase was completely eliminated by holder pasteurization, decreased at 600 MPa, and remained intact at 200 + 400 MPa; 450 MPa. The fatty acid composition and structure of human milk fat triacylglycerols were unchanged. The lipids of human milk after holder pasteurization had the lowest content of free fatty acids and the shortest induction time compared with samples after HPP. HPP slightly changed the β-carotene and lycopene levels, whereas the lutein level was decreased by 40.0% up to 60.2%, compared with 15.8% after the holder pasteurization.

The DGAT1 K232A polymorphism and feeding modify milk fat triacylglycerol composition

In the present study, we aimed to investigate the changes in triacylglycerol (TAG) composition as affected by alterations in the cows' diet due to seasonal variations and genetic factors. For this study, 50 milk fat samples in winter and 50 in summer were used from 25 cows with the DGAT1 KK genotype and 25 cows with the DGAT1 AA genotype. The samples were analyzed for milk fat content (%), fat composition, and TAG composition. We found that the content of TAG species CN54 was higher and that of CN34 and CN36 lower in summer than in winter. This seasonal variation in TAG profile was related to seasonal changes in the fatty acids C14:0, C16:0, C18:0, C18:1 cis-9, total unsaturated fatty acids, and total long-chain fatty acids, most likely resulting from dietary differences between seasons. Furthermore, we quantified the effect of DGAT1 K232A polymorphism on TAG profile and detected a significant effect on TAG species CN36, with higher values for the DGAT1 KK genotype. When adjusting for differences in fat content, we found no significant effects of the DGAT1 K232A polymorphism on TAG profile. We detected a significant interaction between DGAT1 K232A polymorphism and season for TAG species CN42 and CN52; in summer, the KK genotype was associated with higher levels for CN42 than the AA genotype, whereas in winter, the difference between the genotypes was small. For CN52, in summer the AA genotype was associated with higher levels than the KK genotype. In winter, the difference between the genotypes was also small. We show that, regardless of preference for DGAT1 genotype (AA or KK) and depending on the availability of FA according to season, UFA (C18:1 cis-9), short-chain FA (C6:0 and C10:0), and medium-chain FA might be esterified on the glycerol backbone of the TAG, keeping the structure characteristics of each TAG species. To our knowledge, this is the first report on the interaction effect of DGAT1 K232A polymorphism and season on the TAG composition in milk fat.

Inside Front Cover: Research advances based on mass spectrometry for profiling of triacylglycerols in oils and fats and their applications

Inside Front Cover: Research advances based on mass spectrometry for profiling of triacylglycerols in oils and fats and their applications

Research advances based on mass spectrometry for profiling of triacylglycerols in oils and fats and their applications.

Vegetable oils and animal fats are dietary source of lipids that play critical and multiple roles in biological function. Triacylglycerols (TAGs) are the principal component of oils and fats with significant difference in profile among different oils and fats. TAG profiling is essential for nutritional evaluation, quality control and assurance of safety in oils and fats. However, analysis of TAGs is a challenging task because of the complicated composition of TAGs and their similar physicochemical properties in oils and fats. The rapid development of mass spectrometry (MS) technology in recent years makes it possible to analyze the composition, content and structure of TAGs in the study of the physical, chemical and nutritional properties of oils, fats and related products. This review described the research advancement based on MS for profiling of TAGs in oil, fat and their applications in food. The application of MS, including direct infusion strategies, and its combination with chromatography, gas chromatography-MS (GC-MS) and liquid chromatography-MS (LC-MS), in the analysis of TAGs were reviewed. The advantages and disadvantages of these analytical methods with relevant applications for TAGs analysis in food were also described.

Milk fat triacylglycerol packing within processed cheese products

Milk fat triacylglycerol packing within processed cheese products

Milk fat triacylglycerol packing within processed cheese products

Milk fat triacylglycerol packing within processed cheese products

Triacylglycerol regioisomers in human milk resolved with an algorithmic novel electrospray ionization tandem mass spectrometry method

A highly sensitive mass spectrometric (MS) method was developed and validated to analyze ratios of regioisomeric triacylglycerols (TAGs) in fats and oils. UPLC resolution of lithiated TAGs followed by daughter scan MS/MS of positive ions revealed several indicative ions for quantitative analysis. Reference TAGs containing C14–C20 fatty acids (FAs) showed good linear response. Analysis of Finnish and Chinese pooled human milk samples revealed hundreds of regioisomeric TAGs. At least 64mol% of the TAGs were quantified with relative standard deviation <17%. When present in the same TAG molecule together with C18 FAs, palmitic acid was typically in the sn-2 position. When together with FAs 10:0, 12:0, 14:0, 20:1 and 20:2, the sn-2 preference of 16:0 was less clear. Oleic acid occupied typically the sn-1/sn-3 positions but when together with FAs 20:1, 20:2, 18:2, 14:1, 12:0 or 10:0 the positioning of 18:1 did not follow these rules.

Milk fat triacylglycerol packing within processed cheese products

Milk fat triacylglycerol packing within processed cheese products

Milk fat triacylglycerol packing within processed cheese products

Milk fat triacylglycerol packing within processed cheese products