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Lipids represent a large group of biomolecules that are responsible for various functions in organisms. Diseases such as diabetes, chronic inflammation, neurological disorders, or neurodegenerative and cardiovascular diseases can be caused by lipid imbalance. Due to the different stereochemical properties and composition of fatty acyl groups of molecules in most lipid classes, quantification of lipids and development of lipidomic analytical techniques are problematic. Identification of different lipid species from complex matrices is difficult, and therefore individual analytical steps, which include extraction, separation, and detection of lipids, must be chosen properly. This review critically documents recent strategies for lipid analysis from sample pretreatment to instrumental analysis and data interpretation published in the last five years (2019 to 2023). The advantages and disadvantages of various extraction methods are covered. The instrumental analysis step comprises methods for lipid identification and quantification. Mass spectrometry (MS) is the most used technique in lipid analysis, which can be performed by direct infusion MS approach or in combination with suitable separation techniques such as liquid chromatography or gas chromatography. Special attention is also given to the correct evaluation and interpretation of the data obtained from the lipid analyses. Only accurate, precise, robust and reliable analytical strategies are able to bring complex and useful lipidomic information, which may contribute to clarification of some diseases at the molecular level, and may be used as putative biomarkers and/or therapeutic targets. [ABSTRACT FROM AUTHOR]

Acyltransferases (ATs) are important tailoring enzymes that contribute to the diversity of natural products. They catalyze the transfer of acyl groups to the skeleton, which improves the lipid solubility, stability, and pharmacological activity of natural compounds. In recent years, a number of ATs have been isolated from plants. In this review, we have summarized 141 biochemically characterized ATs during the period July 1997 to October 2020, including their function, heterologous expression systems, and catalytic mechanisms. Their catalytic performance and application potential has been further discussed. [ABSTRACT FROM AUTHOR]

The increasing demand for triacylglycerol (TAG) enriching polyunsaturated fatty acids (PUFAs) has led to a surge of interest in microalgal TAG metabolism. Polar membrane lipids serve as the desaturation carrier for PUFA, and the functional group of PUFA can be incorporated into TAG. Monogalactoglycerolipid has been found to provide the de novo synthesized oleate acyl group or the nascent polyunsaturated diacylglycerol backbone for TAG biosynthesis in the model green alga, Chlamydomonas reinhardtii. However, whether other membrane lipids take part in the formation of PUFA-attached TAG has not been clearly discovered. A time course study of glycerolipidomics in the starchless mutant of C. reinhardtii , BAFJ5, which hyper-accumulates TAG, revealed that digalactosyldiacylglycerol (DGDG) and diacylglycerol- N , N , N -trimethylhomoserine (DGTS) turned into the main components of membrane lipids, accounting for 62% of the total polar lipids, under nitrogen deprivation combined with high light conditions. In addition, the membrane lipid molecules DGDG 18:3n3/16:0 and DGTS 16:0/18:3n6 were presumed to be involved in the consecutive integration of the de novo synthesized linolenates into TAG. Based on the stoichiometry calculation, DGDG and DGTS were demonstrated to provide a major contribution to the accumulation of linolenate-attached TAG. Our study gives insights into the potential PUFA-attached TAG formation pathway mediated by the turnover of de novo synthesized DGDG and DGTS in the starchless mutant of Chlamydomonas. [ABSTRACT FROM AUTHOR]

• U. hordei produced mono-acetylated MEL-C as a result of the specificity of UhMat1. • Incorporation of C 16 and C 4 fatty acids in UhMELs depends on enzyme specificities. • Mac1 is responsible for of the small fatty acids at the mannose position C2. • Mac2 adds the medium chain length fatty acids to the mannose residue C3. Mannosylerythritol lipids (MELs) are surface active molecules produced by many basidiomycetous fungi. MELs consist of a mannosylerythritol disaccharide, which is acylated with short and medium chain fatty acids at the mannosyl moiety. A gene cluster composed of five genes is required for MEL biosynthesis. Here we show that the plant pathogenic fungus Ustilago hordei secretes these glycolipids under nitrogen starvation conditions. In contrast to MELs produced by the closely related fungus Ustilago maydis those secreted by U. hordei are mostly mono-acetylated and contain a different mixture of acyl groups. Cross-species complementation between these fungi revealed that these differences result from different catalytic activities of the acetyltransferase Mat1 and the acyltransferases Mac1 and Mac2. U. maydis mat1 mutants expressing the homologous mat1 gene from U. hordei produced mostly mono-acetylated variants and lack di-acetylated MELs normally produced by U. maydis. Furthermore, we determined that the acyltransferase Mac1 acylates the mannosylerythritol moiety at position C2 while Mac2 acylates C3. The identification of decorating enzymes with different substrate specificities will allow the tailor-made production of novel subsets of MELs. [ABSTRACT FROM AUTHOR]

We report a 5-step sequence for the synthesis of phosphatidylglycerols (PG) bearing different or identical chains at positions sn-1 and sn-2 starting from phenyl dichlorophosphate and using (S)-glycidol as the chiral source. Overall, this new approach is more convergent that the previously reported syntheses as the PG derivatives are constructed around the phosphorus center. Hence, this method is more convenient for the incorporation of expensive or complex acyl chains at the sn-2 position. [ABSTRACT FROM AUTHOR]

Oilseed cakes obtained by cold-pressing are rich in lipids susceptible to degradation, which could challenge their handling and nutritional, safety, and sensory requirements, especially during long-storage. Although extrusion is effective at reducing antinutritional factors, its effect on the lipid stability of oilseed cakes remains unknown. In this work, a comparative 1H-NMR-based investigation of the lipid quality in terms of lipid composition, hydrolysis, and oxidation status, as well as their stability after long-term storage was performed on soybean, canola and sunflower cold-pressed cakes before and after low-moisture extrusion. Soybean cake was rich in linolenic and linoleic groups, whereas canola and sunflower mostly contained linoleic and monounsaturated acyl groups and fatty acids. Extrusion did not modify lipid quality immediately after processing. Nevertheless, lipid stability was significantly reduced after 12 months, especially in cakes subjected to extrusion, evidenced by the degradation of their polyunsaturated acyl groups and fatty acids, and the increase of primary and secondary oxidation products. Since lipid quality was retained immediately after extrusion (no storage), results indicated that any action to avoid the enzymatic lipid degradation must be performed immediately after cold-pressing. Moreover, long-term storage should be carefully considered as the lipid stability/quality of extruded cakes was compromised after extrusion. • Lipid quality was studied in terms of acyl group profile, hydrolysis, and oxidation. • Cold-pressed cakes contain up to 18% lipids, most of which susceptible to degradation. • NMR showed that extrusion did not modify lipid quality immediately after processing. • Lipid stability was significantly reduced after 12 months, especially after extrusion. [ABSTRACT FROM AUTHOR]

[Display omitted] • We present an analytical approach for milk polar lipid analysis using UPSFC-MS. • A total of 18 lipid classes were separated and analyzed within 10 min. • 219 lipid molecular species were identified in milk and milk-derived samples. • Polar lipids composition in human milk is obviously different from that in other milk. • Saturation of the acyl groups of different polar lipids in human milk varies considerably. Polar lipids in milk are receiving increasing interest due to their bioactivities. However, milk polar lipids present a wide range of physical–chemical properties at different concentrations, making their analysis challenging. In this study, we presented a comprehensive lipidomic method using ultraperformance supercritical fluid chromatography (UPSFC)-quadrupole-time of flight-mass spectrometry (Q-TOF-MS), which enabled the separation of 18 lipid classes (including nonpolar lipids, cholesterol, ceramide, glycerophospholipids, sphingomyelin, and gangliosides) within 10 min. The method was used to analyze the polar lipids in seven samples, including human milk, other mammalian milk and milk fat globule membrane ingredients, identifying 14 lipid classes containing 219 lipid molecular species. A mass spectrometry data processing strategy applicable for high-throughput studies was also developed and validated. [ABSTRACT FROM AUTHOR]

Headgroup-functionalized lipids arebeing developed as ligand tethers for high selectivity separationson polypropylene capillary-channeled polymer fiber stationary phases.Surface modification is affected under ambient conditions from aqueoussolution. This basic methodology has promise in many areas where robustmodifications are desired on hydrophobic surfaces. In order to understandthe mode of adsorption of the lipid tail to the polypropylene surface,lipids labeled with the environmentally sensitive 7-nitro-2-1,3-benzoxadiazol-4-yl(NBD) fluorophore were used, with NBD covalently attached to the headgroup(NBD-PE) or the acyl chain (acyl NBD-PE) of the lipid. When modifiedwith the acyl NBD-PE, fluorescence imaging of the fiber at excitationwavelengths increasing from 470 to 510 nm caused a 32 nm shift inemission toward the red edge of the absorption band, indicating thatthe NBD molecule (and thus the lipid tail) is motionally restricted.Fluorescence imaging on fibers modified with NBD-PE or the free NBD-Cldye molecule yields no change in the emission response. The resultsof these imaging studies provide evidence that the acyl chain portionsof the lipids intercalate into free volume of the polypropylene fiberstructure, yielding a robust means of surface modification and thepotential for high ligand densities. [ABSTRACT FROM AUTHOR]

TLR4 agonists that favor TRIF-dependent signaling and the induction of type 1 interferons may have potential as vaccine adjuvants with reduced toxicity. CRX-547 ( 4 ), a member of the aminoalkyl glucosaminide 4-phosphate (AGP) class of lipid A mimetics possessing three ( R )-3-decanoyloxytetradecanoyl groups and d -relative configuration in the aglycon, selectively reduces MyD88-dependent signaling resulting in TRIF-selective signaling, whereas the corresponding secondary ether lipid 6a containing ( R )-3-decyloxytetradecanoyl groups does not. In order to determine which secondary acyl groups are important for the reduction in MyD88-dependent signaling activity of 4 , the six possible ester/ether hybrid derivatives of 4 and 6a were synthesized and evaluated for their ability to induce NF-κB in a HEK293 cell reporter assay. An ( R )-3-decanoyloxytetradecanoyl group on the 3-position of the d -glucosamine unit was found to be indispensable for maintaining low NF-κB activity irrespective of the substitutions (decyl or decanoyl) on the other two secondary positions. These results suggest that the carbonyl group of the 3-secondary lipid chain may impede homodimerization and/or conformational changes in the TLR4–MD2 complex necessary for MyD88 binding and pro-inflammatory cytokine induction. [ABSTRACT FROM AUTHOR]

Endotoxin is recognized as one of the virulence factors of the Bordetella avium bird pathogen, and characterization of its structure and corresponding genomic features are important for an understanding of its role in pathogenicity and for an improved general knowledge of Bordetella spp virulence factors. The structure of the biologically active part of B. avium LPS, lipid A, is described and compared to those of another bird pathogen, opportunistic in humans, Bordetella hinzii, and to that of Bordetella trematum, a human pathogen. Sequence analyses showed that the three strains have homologues of acyl-chain modifying enzymes PagL, PagP and LpxO, of the 1-phosphatase LpxE, in addition to LgmA, LgmB and LgmC, which are required for the glucosamine modification. MALDI mass spectrometry identified a high amount of glucosamine substituting the phosphate groups of B. avium lipid A; this modification was absent from B. hinzii and B. trematum. The acylation patterns of the three lipid As were similar, but they differed from those of Bordetella pertussis and Bordetella parapertussis. They were also found to be close to the lipid A structure of Bordetella bronchiseptica, a mammalian pathogen, only differing from the latter by the degree of hydroxylation of the branched fatty acid. [ABSTRACT FROM AUTHOR]

Plant lifespan is affected by factors with genetic and environmental bases. The laws governing these two factors and how they affect plant lifespan are unclear. Here we show that the acyl chain length (ACL) of phosphatidylserine (PS) is correlated with plant lifespan. Among the detected eight head-group classes of membrane lipids with lipidomics based on triple quadrupole tandem mass spectrometry, the ACL of PS showed high diversity, in contrast to the ACLs of the other seven classes, which were highly conserved over all stages of development in all plant species and organs and under all conditions that we studied. Further investigation found that acyl chains of PS lengthened during development, senescence, and under environmental stresses and that increasing length was accelerated by promoted- senescence. The acyl chains of PS were limited to a certain carbon number and ceased to increase in length when plants were close to death. These findings suggest that the ACL of PS can count plant lifespan and could be a molecular scale ruler for measuring plant development and senescence. [ABSTRACT FROM AUTHOR]

We present a new analytical approach for the analysis of triacylglycerol fatty acyls distribution by normal phase liquid chromatography (NPLC) coupled with APPI+-HRMS. The NPLC method used allows the separation of more than 30 classes of lipids. The energy of the APPI+ source enables the formation of low-intensity ions B fragments ([RC = O+74]+ <3%), characteristic of lipids with a glycerol esterified by one or more fatty acyls. We found the relative intensities of ions B were close to the fatty acyl distribution. To establish the proof of concept, we decided to focus on the triacylglycerols (TGs) class, the major component of plant oils. By either NPLC or FIA, the TGs class appeared as a single peak. In our experimental conditions, ions B are always present in the mass spectra of TGs and each ion B is specific to a fatty acyl group. The Orbitrap mass spectrometer featured high enough resolution and accuracy to identify ions B and distinguish them from other TG fragment ions. A further adjustment of the fatty acyls relative quantities calculation from ions B intensities was computed using weighting coefficients of ions B response. The methodology was developed and validated using plant oils characterized by a GC-FID reference method. NPLC-APPI+-HRMS method offers the advantage of analyzing the fatty acyl composition of complex lipid extracts without the need for sample preparation. [Display omitted] • Fatty acyls distribution of triacylglycerols is estimated from their APPI + -HRMS spectra. • Monoacylglycerol like fragments (Ions B) are produced by in source fragmentation of triacylglycerols. • The fatty acyls composition of triacylglycerols (TG) can be deduced from Ion B intensities. • Coupling with flow injection (TG in plant oils) or NPLC (TG in complex samples) is achieved without prior sample preparation. [ABSTRACT FROM AUTHOR]

• Characterization of lipoproteins in plasma is important for cardiovascular risk assessment. • NMR chemical shifts and relaxation time of cholesterol change with pressure. • The T2* of Low Density Lipoprotein (LDL) shows non-linear dependence on pressure. • The discontinuity in T2* of LDL reflect a phase transition at that unique pressure. Determining the particle size and number of lipoprotein components found in blood plasma (HDL, LDL and VLDL) has become an important clinical tool in diagnosing risk of cardiovascular disease. Proton (1H) NMR spectroscopy methods to quantify lipoprotein particle subclasses have been advancing since NMR lineshape analysis of plasma samples was first proposed in the 1990's. NMR methods, including a more recent DOSY-based diffusion spectroscopy test, provide the foundation for the advanced lipoprotein tests, including Lipoprotein® and Liposcale® analyses available for clinical use to determine particle size and number. At the time of this submission, no NMR studies exist which explore physical parameters of individual lipoprotein fractions when they are deformed by pressure. This study reports 1H NMR frequency shifts and T 2 * measurements for the broad methyl peak attributed to terminal methyls (cholesteryl positions 26, 27 and terminal acyl methyl groups) in three primary lipoprotein fractions as a function of hydraulic pressure. This terminal CH 3 resonance shifted linearly upfield as a function of pressure for HDL and VLDL (observed slopes of −0.014 Hz/bar). The LDL terminal CH 3 resonance shows segmented behavior, with a shallow slope between 0–900 bar (−0.008 hz/bar) and a slope similar to HDL and VDL across the range from 1000 to 2400 bar (slope −0.016 Hz/bar). 1H T 2 * values measured for VLDL and HDL dropped linearly with increasing pressure. 1H T 2 * values for LDL demonstrated segmented behavior as a function of pressure. The unique behavior observed for LDL terminal CH 3 frequency and 1H T 2 * trends suggests an approximate pressure at which phase transition occurs. [ABSTRACT FROM AUTHOR]

White adipose tissue (WAT) lipolysis contributes to energy balance during fasting. Lipolysis can proceed by the sequential hydrolysis of triglycerides (TGs) by adipose triglyceride lipase (ATGL), then of diacylglycerols (DGs) by hormone-sensitive lipase (HSL). We showed that the combined genetic deficiency of ATGL and HSL in mouse adipose tissue produces a striking different phenotype from that of isolated ATGL deficiency, inconsistent with the linear model of lipolysis. We hypothesized that the mechanism might be functional redundancy between ATGL and HSL. To test this, the TG hydrolase activity of HSL was measured in WAT. HSL showed TG hydrolase activity. Then, to test ATGL for activity towards DGs, radiolabeled DGs were incubated with HSL-deficient lipid droplet fractions. The content of TG increased, suggesting DG-to-TG synthesis rather than DG hydrolysis. TG synthesis was abolished by a specific ATGL inhibitor, suggesting that ATGL functions as a transacylase when HSL is deficient, transferring an acyl group from one DG to another, forming a TG plus a monoglyceride (MG) that could be hydrolyzed by monoglyceride lipase. These results reveal a previously unknown physiological redundancy between ATGL and HSL, a mechanism for the epistatic interaction between Pnpla2 and Lipe. It provides an alternative lipolytic pathway, potentially important in patients with deficient lipolysis. [ABSTRACT FROM AUTHOR]