Thermophilic bacteria of four genera in contrast to the commonly used production strains such as Bacillus subtilis, produce homologs other than menaquinone (MK) with seven isoprene units. The number of isoprene units and the configuration of double bonds are essential factors for their biological activity. The goal was to obtain a strain of bacteria that produces a wide range of MK homologs and only all-trans geometrical isomers, which was the strain G. kaustophilus. Using off-line two-dimensional LC-tandem MS in columns with the RP18 phase and the COSMOSIL cholester phase (separation according to the geometric configuration of double bonds) it was shown that thermophilic bacteria grown at different temperatures produce only all-trans isomers of menaquinones from MK-5 (menaquinone with five isoprenyl units) to MK-15 (fifteen isoprenyl units). Therefore, G. kaustophilus appears to be a biotechnologically important strain produces only trans isomers and additionally homologs from 5 to 15 isoprene units.
- MeSH
- Bacteria * MeSH
- Butadienes * MeSH
- Mass Spectrometry MeSH
- Vitamin K 2 chemistry MeSH
- Publication type
- Journal Article MeSH
Unusual glucose-substituted cardiolipins (Glcx-CLs) in three genera of thermophilic bacteria, having more than one glycosidically linked glucose to the hydroxyl of the central glycerol of Glcx-CLs were identified for the first time in thermophilic bacteria of the genera Geobacillus, Meiothermus, and Thermus. The number of glucoses reached up to five units. The structure of glycosidically linked oligosaccharides was determined based on shotgun analysis MS (electrospray high-resolution tandem mass spectrometry), partially methylated alditol acetates were identified by GC-MS, both electron ionization (EI) and positive chemical ionization (PCI), hydrophilic interaction liquid chromatography (HILIC) separation and identification of CLs glycosides by high resolution MS-ESI, and digestion by specific glycosidases.
Triacylglycerols (TAGs) containing positional isomers of hypogeic (Hy), palmitoleic (Po), and palmitvaccenic (Pv) acids from three microorganisms (top-fermenting brewer's yeast Saccharomyces cerevisiae, green alga Coccomyxa elongata, and arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis) were analyzed. Dozens of regioisomers and enantiomers of TAGs containing one, two or three hexadecenoic acids have been identified by means of reversed phase chromatography/mass spectrometry (RP-HPLC/MS). The regioisomers of TAGs containing two palmitic acids and any hexadecenoic acid were separated. Analysis of regioisomers of TAGs having one Pv residue showed that asymmetric molecular species such as PvPP or PPPv were dominant in Rhizophagus. TAGs were also analyzed on a chiral phase column and nine molecular species of TAGs containing two palmitic and any of three hexadecenoic acids were separated and identified. In the case of TAGs containing one palmitic and two hexadecenoic acids, the separation was successful only if the hexadecenoic acids were identical. Separation of TAGs containing three hexadecenoic acids was successful only if all three hexadecenoic acids were identical. Regardless of the type of TAG, it was found that TAGs in the AM fungus and containing palmitvaccenic acid bound at the sn-1 position of the glycerol backbone were dominant, suggesting similarity in the biosynthesis of the different TAGs. The covalent adduct chemical ionization method was used for identification of TAGs as adduct with (1-methyleneimino)-1-ethenyl ion, which reacted with double bond of the unsaturated fatty acid. Tandem MS thus makes it possible to identify TAGs containing various hexadecenoic acids.
Very long chain fatty acids (VLCFAs) are important components of various lipid classes in most organisms, from bacteria to higher plants and mammals, including humans. VLCFAs, or very long chain polyunsaturated fatty acids (VLCPUFAs), can be defined as fatty acids with 23 or more carbon atoms in the molecule. The main emphasis in this review is on the analysis of these acids, including obtaining standards from natural sources or their synthesis. Furthermore, the occurrence and analysis of these compounds in both lower (bacteria, invertebrates) and higher organisms (flowering plants or mammals) are discussed in detail. Attention is paid to their biosynthesis, especially the elongation of very long chain fatty acids protein (ELOVL4). This review deals with papers describing these very interesting compounds, whose chemical, biochemical and biological properties have not been fully explored.
Rhamnolipids are extensively studied biosurfactants due to their potential in many industrial applications, eco-friendly production and properties. However, their availability for broader application is severely limited by their production costs, therefore the optimization of efficacy of their cultivation gains significance as well as the information regarding the physio-chemical properties of rhamnolipids resulting from various cultivation strategies. In this work, the bioprocess design focused on optimization of the rhamnolipid yield of Pseudomonas aeruginosa DBM 3774 utilizing the response surface methodology (RSM). Six carbon sources were investigated for their effect on the rhamnolipid production. The RSM prediction improved the total rhamnolipid yield from 2.2 to 13.5 g/L and the rhamnolipid productivity from 11.6 to 45.3 mg/L/h. A significant effect of the carbon source type, concentration and the C/N ratio on the composition of the rhamnolipid congeners has been demonstrated for cultivation of P. aeruginosa DBM 3774 in batch cultivation. Especially, changes in presence of saturated fatty acid in the rhamnolipid congeners, ranging from 18.8% of unsaturated fatty acids (carbon source glycerol; 40 g/L) to 0% (sodium citrate 20 g/L) were observed. This demonstrates possibilities of model based systems as basis in cultivation of industrially important compounds like biosurfactants rhamnolipids and the importance of detailed study of interconnection between cultivation conditions and rhamnolipid mixture composition and properties.
- Publication type
- Journal Article MeSH
A combination of two chromatographic and two enzymatic methods was used for the analysis of molecular species of lipids from Gram-positive bacteria of the genus Kocuria. Gram-positive bacteria contain a majority of branched fatty acids (FAs), especially iso- and/or anteiso-FAs. Two strains K. rhizophila were cultivated at three different temperatures (20, 28, and 37°C) and the majority phospholipid, i.e., the mixture of molecular species of phosphatidylglycerols (PGs) was separated by means of hydrophilic interaction liquid chromatography (HILIC). After enzymatic hydrolysis of PGs by phospholipase C and derivatization of the free OH group, the sn-1,2-diacyl-3-acetyl triacylglycerols (AcTAGs) were separated by reversed phase HPLC. Molecular species such as i-15:0/i-15:0/2:0, ai-15:0/ai-15:0/2:0, and 15:0/15:0/2:0 (straight chains) were identified by liquid chromatography-positive electrospray ionization mass spectrometry. The tandem mass spectra of both standards and natural compounds containing iso, anteiso and straight chain FAs with the same carbons were identical. Therefore, for identification of the ratio of two regioisomers, i.e. i-15:0/ai-15:0/2:0 vs. ai-15:0/i-15:0/2:0, they were cleavage by pancreatic lipase. The mixture of free fatty acids (FFAs) and 2-monoacylglycerols (2-MAGs) was obtained. After their separation by TLC and esterification and/or transesterification, the fatty acid methyl esters were quantified by GC-MS and thus the ratio of regioisomers was determined. It has been shown that the ratio of PG (containing as majority i-15: 0 / i-15: 0, i-15: 0 / ai-15: 0 and / or ai-15: 0 / i-15: 0 and ai-15: 0 / ai-15: 0 molecular species) significantly affected the membrane flow of bacterial cells cultured at different temperatures.
- MeSH
- Chemistry Techniques, Analytical methods MeSH
- Chromatography, Liquid * MeSH
- Diglycerides chemistry isolation & purification MeSH
- Phospholipids chemistry MeSH
- Spectrometry, Mass, Electrospray Ionization * MeSH
- Hydrophobic and Hydrophilic Interactions MeSH
- Fatty Acids chemistry MeSH
- Micrococcaceae chemistry MeSH
- Gas Chromatography-Mass Spectrometry MeSH
- Chromatography, High Pressure Liquid MeSH
- Publication type
- Journal Article MeSH
Background: Hopanoids modify plasma membrane properties in bacteria and are often compared to sterols that modulate membrane fluidity in eukaryotes. In some microorganisms, they can also allow adaptations to extreme environments. Methods: Hopanoids were identified by liquid chromatography-mass spectrometry in fourteen strains of thermophilic bacteria belonging to five genera, i.e., Alicyclobacillus, Brevibacillus, Geobacillus, Meiothermus, and Thermus. The bacteria were cultivated at temperatures from 42 to 70 °C. Results: Regardless of the source of origin, the strains have the same tendency to adapt the hopanoid content depending on the cultivation temperature. In the case of aminopentol, its content increases; aminotetrol does not show a significant change; and in the case of aminotriol the content decreases by almost a third. The content of bacteriohopanetetrol and bacteriohopanetetrol glycoside decreases with increasing temperature, while in the case of adenosylhopane the opposite trend was found. Conclusions: Changes in hopanoid content can be explained by increased biosynthesis, where adenosylhopane is the first intermediate in the biosynthesis of the hopanoid side chain.
- Publication type
- Journal Article MeSH
Plasmalogens are a group of lipids mainly found in the cell membranes. They occur in anaerobic bacteria and in some protozoa, invertebrates and vertebrates, including humans. Their occurrence in plants and fungi is controversial. They can protect cells from damage by reactive oxygen species, protect other phospholipids or lipoprotein particles against oxidative stress, and have been implicated as signaling molecules and modulators of membrane dynamics. Biosynthesis in anaerobic and aerobic organisms occurs by different pathways, and the main biosynthetic pathway in anaerobic bacteria was clarified only this year (2021). Many different analytical techniques have been used for plasmalogen analysis, some of which are detailed below. These can be divided into two groups: shotgun lipidomics, or electrospray ionization mass spectrometry in combination with high performance liquid chromatography (LC-MS). The advantages and limitations of both techniques are discussed here, using examples from anaerobic bacteria to specialized mammalian (human) organs.
Cardiolipins (1,3-bis(sn-3'-phosphatidyl)-sn-glycerol) (CLs) are widespread in many organisms, from bacteria to higher green plants and mammals. CLs were observed in Gram-positive bacterium of the genus Kocuria, brewer's yeast Saccharomyces, the green alga Chlamydomonas, spinach and beef heart. A mixture of molecular species of CLs was obtained from total lipids by hydrophilic interaction liquid chromatography (HILIC), and these were further separated and identified by reversed phase LC/MS with negative tandem electrospray ionization. The majority of CLs molecular species from each organism were cleaved using phospholipase C from Bacillus cereus. This phospholipase cleaves CLs into 1,2-diglycerols and phosphatidylglycerol 3-phosphates, which were then separated. After CLs cleavage, diacylglycerols such as sn-1,2-diacyl-3-acetyl-glycerols (i.e., triacylglycerols) were separated and identified by chiral chromatography/MS-positive tandem ESI. Significant differences in the composition of the molecular species between the 3-(3-sn-phosphatidyl) and 1-(3-sn-phosphatidyl) moieties of CLs were found in all organisms tested. Molecular species of CLs that contained four different fatty acids were identified in all five samples, and CLs containing very long chain fatty acids were identified in yeast. In addition, CLs containing both enantiomers (at the sn-2 carbon) were present in the bacterium tested. These findings were further supported by data already published in GenBank where, in the same family - Micrococcaceae - both enzymes responsible for chirality in the sn-2 position, glycerol-3-phosphate and glycerol-1-phosphate dehydrogenases, were present.
- MeSH
- Chemical Fractionation MeSH
- Chlamydomonas reinhardtii chemistry MeSH
- Chromatography, Liquid methods MeSH
- Spectrometry, Mass, Electrospray Ionization methods MeSH
- Hydrophobic and Hydrophilic Interactions MeSH
- Hydrolysis MeSH
- Cardiolipins chemistry MeSH
- Fatty Acids analysis MeSH
- Cattle MeSH
- Stereoisomerism MeSH
- Triglycerides chemistry MeSH
- Animals MeSH
- Check Tag
- Cattle MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
Polar lipids from the diatoms Diadesmis gallica and Navicula atomus were separated and their structures were determined using high resolution tandem MS HILIC-LC/ESI. This method allowed us to identify 34 classes of lipids, each containing dozens of molecular species, including regioisomers. The largest differences were found in two sulfur-containing lipids, sulfoquinovosyldiacylglycerol and phosphatidylsulfocholine caused probably by the remodeling of lipid species. These diatoms have been found to use several mechanisms to resolve growth in extreme environments, i.e. silica starvation. The presence of insoluble nano-SiO2 leads to the replacement of cellular phospholipids with sulfolipids. Regioisomer ratios also vary depending on the concentration of nano-SiO2 in the culture medium, i.e. the biosynthesis of polar lipids via the prokaryotic (plastidial) and/or eukaryotic (explastidial) pathways. Complex analyses of polar lipids using high resolution HILIC-LC/ESI-tandem, as used for diatoms, can also be used for other photosynthetic microorganisms.
- MeSH
- Spectrometry, Mass, Electrospray Ionization MeSH
- Lipidomics MeSH
- Nanoparticles * MeSH
- Silicon Dioxide MeSH
- Diatoms * MeSH
- Publication type
- Journal Article MeSH
