Cyanobacteria produce a wide range of metabolites of interest for industrial or medical use. The cultivation of freshwater Nostoc cf. linckia yielded 5.4 g/L of a crude exopolysaccharide (cEPS) with a molecular weight of 1.31 × 105 g/mol. Ion-exchange chromatography of cEPS yielded two dominant fractions, EPS-1 and EPS-2, differing in molecular weight. The lower molecular weight fraction (EPS-1) was subjected to structural studies. Results of chemical and spectroscopic analyses showed that three of the four dominant sugars, glucose, galactose and xylose are 1,4-linked in the backbone in the following order: [→4)-β-D-Xylp-(1 → 4)-β-D-Glcp-(1 → 4)-α-D-Galp-(1 → 4)-β-D-Glcp-(1→]n. Terminal mannose residues were identified as side chains linked at C3 of every third backbone xylose and every second glucose is branched at C6 by 3-O-lactyl-β-D-glucuronic acid (nosturonic acid). Antioxidant properties of EPS were tested using two in vitro methods. Both assays showed that the cEPS was more active than purified EPS-1 and EPS-2 fractions and deproteinized EPS.
- MeSH
- antioxidancia chemie MeSH
- bakteriální polysacharidy analýza chemie MeSH
- galaktosa chemie MeSH
- glukosa chemie MeSH
- kyselina glukuronová chemie MeSH
- magnetická rezonanční spektroskopie metody MeSH
- molekulární struktura MeSH
- molekulová hmotnost MeSH
- Nostoc chemie MeSH
- xylosa chemie MeSH
- Publikační typ
- časopisecké články MeSH
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.
Complex structure of cyanobacterium Nostoc sp. exopolysaccharide (EPS), with apparent molecular weight 214 × 103 g/mol, can be deduced from its composition. Chemical and NMR analyses found four dominant sugar monomers, namely (1 → 4)-linked α-l-arabinopyranose, β-d-glucopyranose, β-d-xylopyranose and (1 → 3)-linked β-d-mannopyranose, two different uronic acids and a lactyl group, with (1 → 4,6)-linked β-d-glucopyranose as the only branch point suggest a complex structure of this polymer. The dominant uronic acid is α-linked, but it remained unidentified. β-d-Glucuronic acid was present in lower amount. Their position as well as that of lactyl remained undetermined too. Different doses of orally administered EPS in guinea pigs evoked a significant decrease in cough effort and a decrease in airway reactivity. The antitussive efficacy and bronchodilator effect of higher doses of EPS were found to be similar to that of the antitussive drug codeine and the antiasthmatic salbutamol. Without significant cytotoxicity on the RAW 264.7 cells, EPS stimulated the macrophage cells to produce pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), and prostaglandins (PGs) and nitric oxide (NO) via induction of COX-2 and iNOS expression, respectively, suggesting that this biopolymer potentiates an early innate immune response and can therefore be used as a new immune modulator.
- MeSH
- albuterol farmakologie MeSH
- bakteriální polysacharidy chemie farmakologie MeSH
- biopolymery chemie MeSH
- bronchodilatancia farmakologie MeSH
- buněčné linie MeSH
- cytokiny metabolismus MeSH
- interleukin-6 metabolismus MeSH
- kašel farmakoterapie MeSH
- kodein farmakologie MeSH
- kyselina glukuronová chemie MeSH
- kyseliny uronové chemie MeSH
- makrofágy účinky léků metabolismus MeSH
- morčata MeSH
- myši MeSH
- Nostoc metabolismus MeSH
- oxid dusnatý metabolismus MeSH
- RAW 264.7 buňky MeSH
- sinice metabolismus MeSH
- TNF-alfa metabolismus MeSH
- zvířata MeSH
- Check Tag
- morčata MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Many microalgal species produce a wide range of highly-value products which are interesting for biotechnological applications. Cultivation of microalgal species Dictyosphaerium pulchellum and Dictyosphaerium tetrachotomum, strains Růžicka and Fott resulted yields of 0.2, 0.7 and 1.8 g/L of extracellular biopolymers (EPSs), respectively. All biopolymers were shown to be anionic proteoglycans. The sugar composition analyses of all EPSs showed high contents of hexoses and the presence of partially methylated monosaccharide residues, i.e. hexoses, and deoxy hexoses. The dominant sugar component of all EPSs was found to be galactose. Extracellular microalgal biopolymers were subjected to immunobiological and immunotoxicological evaluation using murine melanoma cancer cells B16, murine fibroblast cell line NIH-3T3, murine macrophages cell line RAW 264.7 and skin construct EpiDerm™ (EPI-200). The EPSs exerted the antiproliferative effectivity; treatment of EPS induced proinflammatory cytokines TNF-α, IL-6, IL-12, IL-1β and IL-17, also engaged in anti-cancer immunity. Immunotoxicological studies revealed their non-toxic character and safe application on EpiDerm™.
- MeSH
- biopolymery chemie farmakologie toxicita MeSH
- buňky NIH 3T3 MeSH
- Chlorophyta chemie MeSH
- extracelulární prostor chemie MeSH
- imunologické faktory chemie farmakologie toxicita MeSH
- kůže účinky léků MeSH
- mikrořasy chemie MeSH
- myši MeSH
- RAW 264.7 buňky MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Genotypic and morphological diversity of cyanobacteria in the Rupite hot spring (Bulgaria) was investigated by means of optical microscopy, cultivation, single-cell PCR, and 16S rRNA gene amplicon sequencing. Altogether, 34 sites were investigated along the 71-39 °C temperature gradient. Analysis of samples from eight representative sites shown that Illumina, optical microscopy, and Roche 454 identified 72, 45 and 19% respective occurrences of all cumulatively present taxa. Optical microscopy failed to detect species of minor occurrence; whereas, amplicon sequencing technologies suffered from failed primer annealing and the presence of species with extensive extracellular polysaccharides production. Amplicon sequencing of the 16S rRNA gene V5-V6 region performed by Illumina identified the cyanobacteria most reliably to the generic level. Nevertheless, only the combined use of optical microscopy, cultivation and sequencing methods allowed for reliable estimate of the cyanobacterial diversity. Here, we show that Rupite hot-spring system hosts one of the richest cyanobacterial flora reported from a single site above 50 °C. Chlorogloeopsis sp. was the most abundant at the highest temperature (68 °C), followed by Leptolyngbya boryana, Thermoleptolyngbya albertanoae, Synechococcus bigranulatus, Oculatella sp., and Desertifilum sp. thriving above 60 °C, while Leptolyngbya geysericola, Geitlerinema splendidum, and Cyanobacterium aponinum were found above 50 °C.
The analysis of triacylglycerols and phospholipids - phosphatidylcholines allowed the use of shotgun lipidomics to identify very long-chain fatty acids and very long-chain polyunsaturated fatty acids in microalgae. These fatty acids were determined in triacylglycerols by positive electrospray ionization of neutral loss scans of different fatty acids, e.g. 24:0, 24:1ω9, 24:6ω3, 26:0, 26:1ω9, 28:0, 28:1ω9, 28:2ω6, and 28:8ω3. Likewise, very long-chain fatty acids in phosphatidylcholines were identified by negative electrospray ionization mass spectrometry in the selected ion-monitoring of the two most important ions (R1COO- and R2COO-). The limit of detection was determined at 10 nmol/L (∼11 pg/μL) in triacylglycerols and 8.6 nmoles/L (∼8 pg/μL) in phosphatidylcholines. The use of liquid chromatography-mass spectrometry is suitable for very long-chain polyunsaturated fatty acids with up to 8 double bonds due to the time of analysis as well as for reasons of lower thermal stability of polyunsaturated fatty acids towards saturated fatty acids, but gas chromatography-mass spectrometry is better suited for the analysis of saturated very long-chain fatty acids.
- MeSH
- Bacteria metabolismus MeSH
- Chlorophyta metabolismus MeSH
- fosfatidylcholiny analýza MeSH
- limita detekce MeSH
- mastné kyseliny analýza MeSH
- nenasycené mastné kyseliny analýza MeSH
- plynová chromatografie s hmotnostně spektrometrickou detekcí MeSH
- referenční standardy MeSH
- triglyceridy analýza MeSH
- Publikační typ
- časopisecké články MeSH
Freshwater bryozoan Pectinatella magnifica was collected from a sand pit (South Bohemia). The total lipids after extraction from lyophilized bryozoans were analyzed using high-performance liquid chromatography/high-resolution negative tandem electrospray mass spectrometry. A total of 19 lipid classes were identified, including N-acyl-substituted phospholipids, that is, N-acylphosphatidylethanolamine and N-acylphosphatidylserine in their plasmenyl forms. Based on gas chromatography/mass spectrometry of 3-pyridylcarbonyl (picolinyl) esters, a very unusual fatty acid was identified, namely 24:7n-3 (all-cis-3,6,9,12,15,18,21-tetracosaheptaenoic acid). The presence of polyunsaturated fatty acids in individual classes is very specific: arachidonic and eicosapentaenoic acids being predominantly bound as amides in N-acyl phospholipids, that is, diacyl-N-acylphosphatidylethanolamines (NAPtdEtn), plasmenyl-N-acylphosphatidyl ethanolamines (PlsNAPtdEtn), diacyl-N-acylphosphatidylserines (NAPtdSer), and plasmenyl-N-acylphosphatidylserines (PlsNAPtdSer). While 24:6n-3 was identified in the sn-2 position of several phospholipids, 24:7n-3 was identified in only two plasmalogens, that is, PlsNAPtdEtn and PlsNAPtdSer. Thanks to the tandem mass spectrometry, we managed to identify the position of all acyl groups in both diacyl- and also in alkenyl-acyl-(plasmenyl) molecular species of N-acylphospholipids. The identification of the molecular species of N-acyl-substituted phosphatidylethanolamine and phosphatidylserine, including their plasmalogen forms, in the freshwater bryozoan P. magnifica has enabled the identification of endogenous cannabinoid precursors.
- MeSH
- antitumorózní látky chemie izolace a purifikace farmakologie MeSH
- biologická dostupnost MeSH
- biopolymery chemie izolace a purifikace farmakologie MeSH
- buněčné linie MeSH
- Chlorophyta chemie MeSH
- cukry chemie izolace a purifikace farmakologie MeSH
- cytokiny metabolismus MeSH
- imunologické faktory chemie izolace a purifikace farmakologie MeSH
- metylace MeSH
- mikrořasy chemie MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- proliferace buněk účinky léků MeSH
- viabilita buněk účinky léků MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Very long chain fatty acids (VLCFAs) were identified in four strains of the green alga Botryococcus braunii (Trebouxiophyceae). The algae contained a series of monoenoic fatty acids up to triacontenoic acid and further VLCFAs in amounts around 1% of total fatty acids. The separation of lipid classes using hydrophilic interaction chromatography revealed that the most abundant VLCFAs (28:2, 28:1 and 28:0) were contained in neutral lipids (triacylglycerols and/or diacylglycerols) and in phospholipids (phosphatidic acid and/or phosphatidylcholine). Using non-aqueous reversed-phase liquid chromatography tandem mass spectrometry (NARP-LC/MS2) of the appropriate collected fractions, molecular species of triacylglycerols containing one or two VLCFAs were described and phosphatidylcholines containing VLCFAs were separated for the first time. Because the presence of Botryosphaerella sudetica (Chlorophyceae) as contaminant of Botryococcus braunii strain Droop 1950/807-1 placed some doubts on the results of previous studies, a strain of this green alga of was also analyzed. In contrast to Botryococcus, C16, a substantially lower proportion of C18 polyunsaturated fatty acids and no VLCFAs were detected in Botryosphaerella.
