Lipidomic and proteomic profiling identifies the milk fat globule membrane composition of milk from cows and camels.

This study was designed to detect lipidomic and proteomic differences in the milk fat globule membrane (MFGM) fractions of cow and camel milk samples. In total, 353 lipid species were detected in these analyses, including 77 PEs, 30 PCs, 28 PIs, 59 SMs, 54 Cers, 13 LPCs, 14 LPEs, 20 PSs, and 4 PGs. These included 54 polar lipid species that differed significantly in abundance between cow and camel milk. Glycerophospholipid metabolism was identified as a core metabolic pathway associated with camel milk composition. Furthermore, 547 proteins exhibiting differential abundance were identified by a label-free proteomics methodology when comparing samples of MFGMfrom camels and cows. Of these proteins, those that differed most in expression between these groups were associated with metabolic pathways, including endoplasmic reticulum activity, endocytosis, and PI3K-Akt signaling. In conclusion, our findings provide a more thorough understanding of the composition of MFGM and its physiological significance, hence offering robust evidence for the potential utilization of camel milk as a nutritional resource in future developments.

Biophysical Interaction Landscape of Mycobacterial Mycolic Acids and Phenolic Glycolipids with Host Macrophage Membranes.

Lipidic adjuvant formulations consisting of immunomodulatory mycobacterial cell wall lipids interact with host cells following administration. The impact of this cross-talk on the host membrane's structure and function is rarely given enough consideration but is imperative to rule out nonspecific perturbation underlying the adjuvant. In this work, we investigated changes in the plasma membranes of live mammalian cells after exposure to mycobacterial mycolic acid (MA) and phenolic glycolipids, two strong candidates for lipidic adjuvant therapy. We found that phenolic glycolipid 1 softened the plasma membrane, lowering membrane tension and stiffness, but MA did not significantly change the membrane characteristics. Further, phenolic glycolipid 1 had a fluidizing impact on the host plasma membrane, increasing the fluidity and the abundance of fluid-ordered-disordered coexisting lipid domains. Notably, lipid diffusion was not impacted. Overall, MA and, to a lesser extent, phenolic glycolipid 1, due to minor disruption of host cell membranes, may serve as appropriate lipids in adjuvant formulations.

Independent and interactive associations between greenness and ambient pollutants on novel glycolipid metabolism biomarkers: A national repeated measurement study.

This study aims to investigate the independent and interactive effects of greenness and ambient pollutants on novel glycolipid metabolism biomarkers. A repeated national cohort study was conducted among 5085 adults from 150 counties/districts across China, with levels of novel glycolipid metabolism biomarkers of TyG index, TG/HDL-c, TC/HDL-c, and non-HDL-c measured. Exposure levels of greenness and ambient pollutants (including PM1, PM2.5, PM10, and NO2) for each participant were determined based on their residential location. Linear mixed-effect and interactive models were used to evaluate the independent and interactive effects between greenness and ambient pollutants on the four novel glycolipid metabolism biomarkers. In the main models, the changes [ß (95% CIs)] of TyG index, TG/HDL-c, TC/HDL-c, and non-HDL-c were -0.021 (-0.036, -0.007), -0.120 (-0.175, -0.066), -0.092 (-0.122, -0.062), and -0.445 (-1.370, 0.480) for every 0.1 increase in NDVI, and were 0.004 (0.003, 0.005), 0.014 (0.009, 0.019), 0.009 (0.006, 0.011), and 0.067 (-0.019, 0.154) for every 1 µg/m3 increase in PM1. Results of interactive analyses demonstrated that individuals living in low-polluted areas could get greater benefits from greenness than those living in highly-polluted areas. Additionally, the results of mediation analyses revealed that PM2.5 mediated 14.40% of the association between greenness and the TyG index. Further research is needed to validate our findings.

A novel bacteriobiont of the Arctic lichen Flavocetraria nivalis, Lichenifustis flavocetrariae gen. nov, sp. nov. demonstrating hydrolytic properties and containing a full set of the Calvin-Benson-Bassham cycle genes.

A Gram-negative, strictly aerobic, chemoorganotrophic, bacteriochlorophyll a-containing, slow-growing bacterium was isolated from the lichen Flavocetraria nivalis and designated strain BP6-180914 T. Cells of this strain were large nonmotile rods, which reproduced by binary fission. Cells grew under oxic conditions and were able to utilize sugars and several polysaccharides, including starch and pectin. Strain BP6-180914 T was psychrotolerant and moderately acidophilic growing at 4-35 °C (optimum 20-28 °C) and between pH 4.0 and 7.5 (optimum 4.5-5.5). The major fatty acids were C18:1ω7c, C19:0 cyclo, C16:0 and C18:0. The polar lipids were diphosphatidylglycerols, phosphatidylglycerols, phosphatidylethanolamines, phosphatidylcholines, unidentified aminolipids, and a number of glycolipids, the major one being an unidentified glycolipid. The quinone was Q-10. The DNA G + C content was 63.65%. Comparative 16S rRNA gene sequence analysis revealed that strain BP6-180914 T was a member of the order Hyphomicrobiales and belonged to the family Lichenihabitantaceae defined by the lichen-dwelling facultative aerobic chemo-organotroph Lichenihabitans psoromatis (92.7% sequence similarity). The results of phylogenomic and genomic relatedness analyses showed that strain BP6-180914 T could clearly be distinguished from other species in the order Hyphomicrobiales with average nucleotide identity values of < 74.05% and genome-to-genome distance values of < 21.1%. The AAI value of 65.9% between strain BP6-180914 T and L. psoromatis allowed us to assign this strain to the novel genus of the family Lichenihabitantaceae. Therefore, it is proposed that strain BP6-180914 T represents a novel species in a new genus, Lichenifustis flavocetrariae gen. nov., sp. nov.; strain BP6-180914 T (= KCTC 92872 T = VKM B-3641 T = UQM 41506 T) is the type strain.

Structural Analyses of the Glycolipids in Lipid Rafts.

Lipid rafts are usually isolated from cells or tissues using sucrose gradient ultracentrifugation in the presence of detergents such as Triton X-100 at 4 °C. Although detergents should be removed for further structural characterization following fractionation, these compounds are often difficult to completely remove, especially from the glycolipids. In this chapter, we describe a novel method for the fast and convenient removal of detergents from lipid raft glycolipids following fraction and describe the application of this method.

NMR Analysis of Mammalian Glycolipids.

Mammalian glycolipids play a variety of roles, often coupled with interactions with endogenous and exogenous molecules. The interactions can induce intracellular signaling and are the means by which glycolipids express biological phenotypes. Insights into the structure-function relationships of glycolipids (both glycan and lipid moieties) provide the basis for gaining an understanding of the mechanisms at play, an important area for further study. Solution nuclear magnetic resonance (NMR) spectroscopy is a unique and powerful method employed to provide, at the atomic level, structural information on glycolipids and other biomolecules in solutions. This chapter briefly describes how we measure NMR spectra of glycolipids and the information gained from NMR spectral analysis.

Deinococcus betulae sp. nov. and Deinococcus arboris sp. nov., novel bacteria isolated from bark of birch tree (Betula platyphylla).

Two reddish-coloured bacterial strains (HMF7604T and HMF7620T) were isolated from bark of birch tree (Betula platyphylla) together with two strains (designed as HMF7603 and HMF7618). Cells were observed to be Gram-stain-negative, oval- to short rod-shaped and non-motile. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the four isolates belonged to the genus Deinococcus, family Deinococcaceae. They had the highest similarities (95.4-95.6 %) to Deinococcus multiflagellatus ID1504T, with which they formed a clade in phylogenetic trees. Menaquinone-8 was the only respiratory quinone. The predominant fatty acids were summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c), C15 : 1 ω6c, C17 : 0 and C16 : 0. Strain HMF7604T contained two unidentified phosphoglycolipids, nine unidentified glycolipids, one unidentified aminolipid, three unidentified phospholipids and three unidentified polar lipids, while strain HMF7620T contained one unidentified phosphoglycolipid, four unidentified glycolipids, one unidentified aminophospholipid, one unidentified phospholipid and one unidentified polar lipid. The DNA G+C contents of strains HMF7604T and HMF7620T were 65.6 and 65.7 mol%, respectively. The average nucleotide identity and digital DNA-DNA hybridization values between the two isolates and their close relative D. multiflagellatus were 81.1-95.3 and 24.5-61.6 %, respectively. Based on the results of phenotypic and phylogenetic characterizations, the four isolates are considered to represent two novel species of the genus Deinococcus, for which the names Deinococcus betulae sp. nov. and Deinococcus arboris sp. nov. are proposed. The type strains are HMF7604T (=KCTC 43354T=NBRC 115489T) and HMF7620T (=KCTC 43051T=NBRC 113959T).

Lysobacter selenitireducens sp. nov., isolated from river sediment.

A Gram-stain-negative, yellow-pigmented, motile, flagellated and rod-shaped bacterium, designated as 13AT, was isolated from a river sediment sample of Fuyang River in Hengshui City, Hebei Province, PR China. Strain 13AT grew at 10-37 °C (optimum, 30 °C), at pH 5.0-11.0 (optimum, pH 7.0) and at 0-7 % (w/v) NaCl concentration (optimum, 0 %). Phylogenetic analysis based on the 16S rRNA gene sequence showed that strain 13AT belongs to the genus Lysobacter, and was most closely related to Lysobacter spongiicola DSM 21749T (97.8 %), Lysobacter concretionis DSM 16239T (97.5 %), Lysobacter daejeonensis GIM 1.690T (97.3 %) and Lysobacter arseniciresistens CGMCC 1.10752T (96.9 %). Meanwhile, the type species Lysobacter enzymogenes ATCC 29487T was selected as a reference strain (95.2 %). The genomic size of strain 13AT was 3.0 Mb and the DNA G+C content was 69.0 %. The average nucleotide identity values between strain 13AT and each of the reference type strains L. spongiicola DSM 21749T, L. concretionis DSM 16239T, L. daejeonensis GIM 1.690T, L. arseniciresistens CGMCC 1.10752T and L. enzymogenes ATCC 29487T were 75.9, 76.1, 77.7, 78.0 and 73.2 %, respectively. The digital DNA-DNA hybridization values between strain 13AT and each of the reference type strains were 21.7, 22.2, 21.9, 22.7 and 23.2 %, respectively. The average amino acid identity values between strain 13AT and each of the reference type strains were 72.5, 72.9, 72.3, 75.0 and 69.2 %, respectively. The major fatty acids were iso-C15 : 0, iso-C16 : 0 and summed feature 9 (iso-C17 : 1 ω9c and/or C16 : 0 10-methyl). The sole respiratory quinone was identified as ubiquinone-8. The polar lipid profile contained phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, an unidentified aminolipid, an unidentified lipid, four unidentified phospholipids and two unidentified glycolipids. Based on the phenotypic, physiological, phylogenetic and chemotaxonomic data, strain 13AT represents a novel species of the genus Lysobacter, for which the name Lysobacter selenitireducens sp. nov. is proposed. The type strain is 13AT (=JCM 34786T=GDMCC 1.2722T).

Sphingomicrobium nitratireducens sp. nov., isolated from a tidal flat in Guangxi.

An aerobic, yellow-pigmented and Gram-stain-negative strain, designated as O-35 T, was isolated from a tidal flat sediment collected in Dangjiang Town, the southern China. Colonies of strain O-35 T were circular with 0.5-1.0 mm in diameter, convex and smooth. Cells of strain O-35 T were coccoid-shaped, non-spore forming, non-motile and the strain could reduce nitrate. Growth of strain O-35 T was observed at 15-40 °C (optimum 30 °C), at pH 6.0-9.5 (optimum 7.5-8.0) and in 0.5-5.0% NaCl (optimum 2%, w/v). Strain O-35 T showed 16S rRNA gene sequence identities of 97.3-97.5% with Sphingomicrobium lutaoense CC-TBT-3 T and Sphingomicrobium aestuariivivum AH-M8T, higher than the rest of Sphingomicrobium type strains. Phylogenetic trees based on the 16S rRNA gene and the core-genome sequences demonstrated that strain O-35 T was affiliated within the genus Sphingomicrobium. Overall genome relatedness index calculations revealed that strain O-35 T had < 75.8% of average nucleotide identity and < 19.2% of digital DNA-DNA hybridization values with Sphingomicrobium type strains. The sole isoprenoid quinone was ubiquinone-10. The major fatty acids (> 10%) were summed feature 8, summed feature 3, C16:0 and C18:1 2-OH. The polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, sphingoglycolipid, two unidentified glycolipids, one unidentified lipid and one unidentified phospholipid. On the basis of the phenotypic, chemotaxonomic and genomic properties, strain O-35 T represents a novel species in the genus Sphingomicrobium, for which the name Sphingomicrobium nitratireducens sp. nov. is proposed. The type strain is O-35 T (= KCTC 92308 T = MCCC 1K07589T).

Deinococcus taeanensis sp. nov., a Radiation-Resistant Bacterium Isolated from a Coastal Dune.

A Gram-stain-negative, nonspore-forming, nonmotile, aerobic, rod-shaped, and very pale orange-colored bacterial strain, designated TS293T, was isolated from a sand sample obtained from a coastal dune after exposure to 3kGy of gamma (γ)-radiation. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that the isolate was a member of the genus Deinococcus and clustered with D. deserti VCD115T. The genome of strain TS293T was 4.62 Mbp long (68.2% G + C content and 4124 predicted genes) divided into a 2.86Mb main chromosome and five plasmids. Many genes considered to be important to the γ-radiation and oxidative stress resistance of Deinococcus were conserved in TS293T, but genome features that could differentiate TS293T from D. deserti and D. radiodurans, the type species of the Deinococcus genus, were also detected. Strain TS293T showed resistance to γ-radiation with D10 values (i.e., the dose required to reduce the bacterial population by tenfold) of 3.1kGy. The predominant fatty acids of strain TS293T were summed feature 3 (C16:1 ω6c and/or C16:1 ω7c) and iso-C16:0. The major polar lipids were two unidentified phosphoglycolipids and one unidentified glycolipid. The main respiratory quinone was menaquinone-8. Based on the phylogenetic, genomic, physiological, and chemotaxonomic characteristics, strain TS293T represents a novel species, for which the name Deinococcus taeanensis sp. nov. is proposed. The type strain is TS293T (= KCTC 43191T = JCM 34027T).

Saccharothrix obliqua sp. nov., isolated from soil of Sichang Island, Thailand.

During an investigation of rare actinobacteria, isolate SC076T was isolated from a soil sample collected from Sichang Island, Chonburi Province, Thailand. The strain showed the highest 16S rRNA gene similarity to Saccharothrix australiensis DSM 43800T (98.6%) and Saccharothrix espanaensis DSM 44229T (98.6%). The zigzag morphology of the spore chain was observed on the aerial mycelia. meso-Diaminopimelic acid was detected in the peptidoglycan. Whole-cell sugars contained rhamnose, ribose, mannose glucose and galactose. Polar lipids were phosphatidylethanolamine, hydroxyphosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol mannoside, phosphatidylinositol, unidentified ninhydrin-positive glycolipid, unidentified glycolipid and four unidentified lipids. The menaquinones were MK-9(H8), MK-9(H4), MK-9(H2) and MK-9(H0). The predominant fatty acids were iso-C16 : 0, iso-C15 : 0 and anteiso-C17 : 0. The draft genome of SC076T was 8040245 bp with a G+C content of 72.5 mol%. The results of genomic analysis between strain SC076T and the related type strains showed that the digital DNA-DNA hybridization and average nucleotide identity values among the strains were 23.6-32.8% and 77.7-86.8 %, respectively, which are lower than the thresholds used to distinguish strains from others of the same species. Based on the taxonomic evidence, strain SC076T represents a novel species of the genus Saccharothrix for which the name Saccharothrix obliqua sp. nov. is proposed. The type strain is SC076T (=TBRC 14540T=NBRC 115117T).

The lipid composition of the in situ pellicle.

OBJECTIVE: The present study aimed to systematically analyse the complete lipid profile of the in situ pellicle in comparison to saliva. For the first time, the modern sensitive methods GC-EI/MS and HPLC MS/MS were to be used for this purpose. DESIGN: Bovine enamel slabs were exposed to the oral cavity of 12 subjects by customized splints (3 min, 30 min or 120 min). Afterwards, the pellicle samples were obtained and further investigated in vitro. Additionally, corresponding unstimulated saliva samples were collected. GC-EI/MS was performed to qualitatively and quantitatively determine all fatty acids contained in the investigated samples. The individual lipid classes of phospholipids, triacylglycerols, glycolipids, cholesterol and cholesterol esters were analysed qualitatively by HPLC MS/MS. RESULTS: A characteristic fatty acid profile of the in situ pellicle was proven. Furthermore, triacylglycerols with the major fatty acids 16:0, 18:0, 18:1, 18:2, and phospholipids were detected as integral components in the pellicle. There were four groups of phospholipids: Lyso-phosphatidylcholines, phosphatidylcholines, phosphatidylethanol-amines, and phosphatidylinositols. Differences between saliva and pellicle were evident in the composition of the fatty acid- and the phospholipid profile. Glycolipids, cholesterol and cholesterol esters could neither be detected in pellicle- nor in saliva samples. CONCLUSION: The lipid profiles of the in situ pellicle and saliva were successfully characterised. Differences in the phospholipid and fatty acid composition between pellicle and saliva indicate a selective pellicle formation process. The results provide an important reference and core data for further investigation of the complex surface interactions in the oral cavity, especially concerning hydrophobic substances.

Halocatena salina sp. nov., a filamentous halophilic archaeon isolated from Aiding Salt Lake.

A filamentous cell-shaped halophilic archaeon (strain AD-1T) was isolated from Aiding Salt Lake, PR China. Its colonies on HCM7 agar plates were pinkish white, 1-4 mm (diameter), elevated and round. The optimum conditions for growth were observed at 42 °C, 4.3 M NaCl, 0.01 M MgCl2 and pH 7. Strain AD-1T could hydrolyse Tween 60, Tween 80, starch and gelatin. Phylogenetic analysis based on 16S rRNA gene, rpoB' and the concatenated 484 single-copy orthologous proteins revealed that strain AD-1T formed a clade with Halocatena pleomorpha SPP-AMP-1T. The average nucleotide identity and in silico DNA-DNA hybridization values between strain AD-1T and Halocatena pleomorpha SPP-AMP-1T were both below the species delineation thresholds (95~96 and 70 %, respectively). The major phospholipids of strain AD-1T were phosphatidic acid, phosphatidylglycerol and phosphatidylglycerol phosphate methyl ester, while the major glycolipids were sulphated galactosyl mannosyl glucosyl diether, galactosyl mannosyl glucosyl diether and glucosyl mannosyl glucosyl diether. The phenotypic, phylogenetic and genome-based analyses suggested that strain AD-1T (=CGMCC 1.13724T=JCM 32960T) represents a novel species, for which the name Halocatena salina sp. nov. is proposed.

Detection of Bacterial Rhamnolipid Toxin by Redox Liposome Single Impact Electrochemistry.

The detection of Rhamnolipid virulence factor produced by Pseudomonas aeruginosa involved in nosocomial infections is reported by using the redox liposome single impact electrochemistry. Redox liposomes based on 1,2-dimyristoyl-sn-glycero-3-phosphocholine as a pure phospholipid and potassium ferrocyanide as an encapsulated redox content are designed for using the interaction of the target toxin with the lipid membrane as a sensing strategy. The electrochemical sensing principle is based on the weakening of the liposomes lipid membrane upon interaction with Rhamnolipid toxin which leads upon impact at an ultramicroelectrode to the breakdown of the liposomes and the release/electrolysis of its encapsulated redox probe. We present as a proof of concept the sensitive and fast sensing of a submicromolar concentration of Rhamnolipid which is detected after less than 30 minutes of incubation with the liposomes, by the appearing of current spikes in the chronoamperometry measurement.

Healthy Breastfeeding Infants Consume Different Quantities of Milk Fat Globule Membrane Lipids.

The human milk fat globule membrane (MFGM) contains important lipids for growing infants. Anthropometric measurements, milk samples, and infant milk intake were collected in a cohort of eleven healthy mother-infant dyads during exclusive breastfeeding from birth to six months. One hundred and sixty-six MFGM lipids were analysed using liquid chromatography-mass spectrometry, and the infant intake was calculated. The concentrations and intake were compared and associations between infant intake and growth characteristics explored. The lipid concentrations and infant intake varied widely between mother-infant dyads and between months one and three. The infant intake for many species displayed positive correlations with infant growth, particularly phospholipid species. The high variation in lipid intake is likely an important factor in infant growth, with strong correlations identified between the intake of many MFGM lipids and infant head circumference and weight. This study highlights the need for intake measurements and inclusion in cohort studies to elucidate the role of the human milk lipidome in infant growth and development.

Potential Benefits of Bovine Colostrum in Pediatric Nutrition and Health.

Bovine colostrum (BC), the first milk produced from cows after parturition, is increasingly used as a nutritional supplement to promote gut function and health in other species, including humans. The high levels of whey and casein proteins, immunoglobulins (Igs), and other milk bioactives in BC are adapted to meet the needs of newborn calves. However, BC supplementation may improve health outcomes across other species, especially when immune and gut functions are immature in early life. We provide a review of BC composition and its effects in infants and children in health and selected diseases (diarrhea, infection, growth-failure, preterm birth, necrotizing enterocolitis (NEC), short-bowel syndrome, and mucositis). Human trials and animal studies (mainly in piglets) are reviewed to assess the scientific evidence of whether BC is a safe and effective antimicrobial and immunomodulatory nutritional supplement that reduces clinical complications related to preterm birth, infections, and gut disorders. Studies in infants and animals suggest that BC should be supplemented at an optimal age, time, and level to be both safe and effective. Exclusive BC feeding is not recommended for infants because of nutritional imbalances relative to human milk. On the other hand, adverse effects, including allergies and intolerance, appear unlikely when BC is provided as a supplement within normal nutrition guidelines for infants and children. Larger clinical trials in infant populations are needed to provide more evidence of health benefits when patients are supplemented with BC in addition to human milk or formula. Igs and other bioactive factors in BC may work in synergy, making it critical to preserve bioactivity with gentle processing and pasteurization methods. BC has the potential to become a safe and effective nutritional supplement for several pediatric subpopulations.

Lipidomics of the Edible Brown Alga Wakame (Undaria pinnatifida) by Liquid Chromatography Coupled to Electrospray Ionization and Tandem Mass Spectrometry.

The lipidome of a brown seaweed commonly known as wakame (Undaria pinnatifida), which is grown and consumed around the world, including Western countries, as a healthy nutraceutical food or supplement, was here extensively examined. The study was focused on the characterization of phospholipids (PL) and glycolipids (GL) by liquid chromatography (LC), either hydrophilic interaction LC (HILIC) or reversed-phase LC (RPLC), coupled to electrospray ionization (ESI) and mass spectrometry (MS), operated both in high and in low-resolution mode. Through the acquisition of single (MS) and tandem (MS/MS) mass spectra more than 200 PL and GL of U. pinnatifida extracts were characterized in terms of lipid class, fatty acyl (FA) chain composition (length and number of unsaturations), and regiochemistry, namely 16 SQDG, 6 SQMG, 12 DGDG, 5 DGMG, 29 PG, 8 LPG, 19 PI, 14 PA, 19 PE, 8 PE, 38 PC, and 27 LPC. The FA (C16:0) was the most abundant saturated acyl chain, whereas the monounsaturated C18:1 and the polyunsaturated C18:2 and C20:4 chains were the prevailing ones. Odd-numbered acyl chains, iJ., C15:0, C17:0, C19:0, and C19:1, were also recognized. While SQDG exhibited the longest and most unsaturated acyl chains, C18:1, C18:2, and C18:3, in the sn-1 position of glycerol, they were preferentially located in the sn-2 position in the case of PL. The developed analytical approach might pave the way to extend lipidomic investigations also for other edible marine algae, thus emphasizing their potential role as a source of bioactive lipids.

Surface-assisted Laser Desorption/ionization Mass Spectrometry Analysis of the Glycolipid Biosurfactants, Mannosylerythritol Lipids, Using an Ionization-assisting Substrate.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a promising tool for the screening of glycolipid-type biosurfactants (BSs) from a crude extract of microbial products. However, it is unsuitable for the detection of lower molecular weight products because the observed ions are overlapped with matrix-derived ions at lower mass range. In this study, we applied a "matrix-free" surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) analysis using a through-hole alumina membrane as an ionization-assisting substrate. Using this method, we could detect a variety of lower molecular weight products in an extract of a glycolipid BS producer with good sensitivity. In addition, the culture solution could be analyzed directly by this method.

Targeted Lipidomics of Mycobacterial Lipids and Glycolipids.

Decades of study have highlighted the richness and uniqueness of the repertoire of lipid and glycolipid families produced by mycobacteria. Many of these families potently regulate host immune responses, in stimulatory or suppressive ways. Thus, the global study of this repertoire in different genetic backgrounds or under model conditions of infection is gaining interest. Despite the difficulties associated with the specificities of this repertoire, the field of mass spectrometry-based lipidomics of mycobacteria has recently made considerable progress, particularly at the analytical level. There is still considerable scope for further progress, especially with regard to the development of an efficient bioinfomatics pipeline for the analysis of the large datasets generated. This chapter describes an HPLC-MS methodology allowing the simultaneous screening of more than 20 of the lipid families produced by mycobacteria and provides recommendations to analyze the generated data given the state-of-the-art.

Characterization of Glucuronosyl-diacyl/monoacylglycerols and Discovery of Their Acylated Derivatives in Tomato Lipid Extracts by Reversed-Phase Liquid Chromatography with Electrospray Ionization and Tandem Mass Spectrometry.

Glucuronic acid containing diacylglycerols (3-(O-α-d-glucuronopyranosyl)-1,2-diacyl-sn-glycerols, GlcA-DAG) are glycolipids of plant membranes especially formed under phosphate-depletion conditions. An analytical approach for the structural characterization of GlcA-DAG in red ripe tomato (Solanum lycopersicum L.) extracts, based on reversed-phase liquid chromatography (RPLC) coupled with electrospray ionization (ESI) and tandem mass spectrometry (MS/MS) using a linear ion trap, is described in this paper. At least 14 GlcA-DAG (R1/R2) species, including four regioisomers, containing three predominant fatty acyl chains C16:0, C18:2, and C18:3, were identified for the first time. Moreover, 29 GlcA-DAG acylated on the glucuronosyl ring (acyl-R3 GlcA-DAG) were discovered, alongside 15 acylated lyso-forms, i.e., acylated 3-(O-α-d-glucuronosyl)monoacylglycerols, abbreviated as acyl-R3 GlcA-MAG (R1/0) or (0/R2). Although many of these acylated lyso-forms were isomeric with GlcA-DAG (i.e., acyl chains with equivalent sum composition), they were successfully separated by reversed-phase liquid chromatography (RPLC) using a solid-core C18 column packed with 2.6 µm particle size. Tandem MS (and eventually MS3) data obtained from sodium adducts ([M + Na]+) and deprotonated molecules ([M - H]-) were fundamental to detect diagnostic product ions related to the glucuronosyl ring and then determine the identity of all investigated glycolipids, especially to recognize the acyl chain linked to the ring. A classification of GlcA-MAG, GlcA-DAG, and acylated GlcA-DAG and GlcA-MAG was generated by an in house-built database. The discovery of acylated derivatives emphasized the already surprising heterogeneity of glucuronic acid-containing mono- and diacylglycerols in tomato plants, stimulating interesting questions on the role played by these glycolipids.