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The structure of lipid A from lipopolysaccharide (LPS) of Rhodomicrobium vannielii ATCC 17100 (Rv) a phototrophic, budding bacterium was re-investigated using high-resolution mass spectrometry, NMR, and chemical degradation protocols. It was found that the (GlcpN)-disaccharide lipid A backbone was substituted by a GalpA residue that was connected to C-1 of proximal GlcpN. Some of this GalpA residue was &beta, eliminated by alkaline de-acylation, which indicated the possibility of the presence of another so far unidentified substituent at C-4 in non-stoichiometric amounts. One Manp residue substituted C-4&prime, of distal GlcpN. The lipid A backbone was acylated by 16:0(3-OH) at C-2 of proximal GlcpN, and by 16:0(3-OH), i17:0(3-OH), or 18:0(3-OH) at C-2&prime, of distal GlcpN. Two acyloxy-acyl moieties that were mainly formed by 14:0(3-O-14:0) and 16:0(3-O-22:1) occupied the distal GlcpN of lipid A. Genes that were possibly involved in the modification of Rv lipid A were compared with bacterial genes of known function. The biological activity was tested at the model of human mononuclear cells (MNC), showing that Rv lipid A alone does not significantly stimulate MNC. At low concentrations of toxic Escherichia coli O111:B4 LPS, pre-incubation with Rv lipid A resulted in a substantial reduction of activity, but, when higher concentrations of E. coli LPS were used, the stimulatory effect was increased.

The free-living Gram-negative bacterium Oligotropha carboxidovorans (formerly: Pseudomonas carboxydovorans), isolated from wastewater, is able to live in aerobic and, facultatively, in autotrophic conditions, utilizing carbon monoxide or hydrogen as a source of energy. The structure of O. carboxidovorans lipid A, a hydrophobic part of lipopolysaccharide, was studied using NMR spectroscopy and high-resolution mass spectrometry (MALDI-ToF MS) techniques. It was demonstrated that the lipid A backbone is composed of two d-GlcpN3N residues connected by a β-(1→6) glycosidic linkage, substituted by galacturonic acids (d-GalpA) at C-1 and C-4’ positions. Both diaminosugars are symmetrically substituted by 3-hydroxy fatty acids (12:0(3-OH) and 18:0(3-OH)). Ester-linked secondary acyl residues (i.e., 18:0, and 26:0(25-OH) and a small amount of 28:0(27-OH)) are located in the distal part of lipid A. These very long-chain hydroxylated fatty acids (VLCFAs) were found to be almost totally esterified at the (ω-1)-OH position with malic acid. Similarities between the lipid A of O. carboxidovorans and Mesorhizobium loti, Rhizobium leguminosarum, Caulobacter crescentus as well as Aquifex pyrophylus were observed and discussed from the perspective of the genomic context of these bacteria.

Exopolymeric substances (EPS) can determine plant-microorganism interactions and have great potential as bioactive compounds. The different amounts of EPS obtained from cultures of three endophytic Fusarium culmorum strains with different aggressiveness&mdash, growth promoting (PGPF), deleterious (DRMO), and pathogenic towards cereal plants&mdash, depended on growth conditions. The EPS concentrations (under optimized culture conditions) were the lowest (0.2 g/L) in the PGPF, about three times higher in the DRMO, and five times higher in the pathogen culture. The EPS of these strains differed in the content of proteins, phenolic components, total sugars, glycosidic linkages, and sugar composition (glucose, mannose, galactose, and smaller quantities of arabinose, galactosamine, and glucosamine). The pathogen EPS exhibited the highest total sugar and mannose concentration. FTIR analysis confirmed the &beta, configuration of the sugars. The EPS differed in the number and weight of polysaccharidic subfractions. The EPS of PGPF and DRMO had two subfractions and the pathogen EPS exhibited a subfraction with the lowest weight (5 kDa). The three EPS preparations (ethanol-precipitated EP, crude C, and proteolysed P) had antioxidant activity (particularly high for the EP-EPS soluble in high concentrations). The EP-EPS of the PGPF strain had the highest antioxidant activity, most likely associated with the highest content of phenolic compounds in this EPS.

Wild mushrooms are gathered and consumed in many regions of the world. Edible mushrooms are a good source of valuable nutritious ingredients but they also are able to accumulate heavy metals from the environment. Participation of polysaccharide components of mushroom cell walls in biosorption is poorly understood, therefore, our investigations focussed on an alkali-soluble polysaccharide derived from the cell wall of Boletus edulis. Its chemical structure is described based on chemical analysis, GPC, NMR and FT-IR and other instrumental techniques. The polysaccharide contained mainly C-3 substituted α- d -glucose and α- d -mannose. The ratio between these components was 10:4. We postulate the polymer is a (1 → 3)-linked α- d -glucan decorated by α-(1 → 3)- d -mannose chains. The molecular mass of the polysaccharide was estimated at about 850 kDa. Biosorption of heavy metal ions (Cd, Cu, Zn and Pb) by the tested polymer was measured using AAS techniques. The polysaccharide has a high ability to accumulate lead and cadmium.

The structure of lipid A from lipopolysaccharide of Phyllobacterium trifolii PETP02T, a nitrogen-fixing symbiotic bacterium, was studied. It was found that the lipid A backbone was composed of two 2,3-diamino-2,3-dideoxy-d-glucose (GlcpN3N) residues connected by a β-(1 → 6) glycosidic linkage, substituted by galacturonic acid (GalpA) at position C-1 and partly decorated by a phosphate residue at C-4′ of the non-reducing GlcpN3N. Both diaminosugars were symmetrically substituted by 3-hydroxy fatty acids (14:0(3-OH) and 16:0(3-OH)). Ester-linked secondary acyl residues [i.e. 19:0cyc and 28:0(27-OH) or 28:0(27-4:0(3-OMe))] were located in the distal part of lipid A. A high similarity between the lipid A of P. trifolii and Mesorhizobium was observed and discussed from the perspective of the genetic context of both genomes.

This review focuses on the chemistry and structures of (Brady)rhizobium lipids A, indispensable parts of lipopolysaccharides. These lipids contain unusual (ω-1) hydroxylated very long chain fatty acids, which are synthesized by a very limited group of bacteria, besides rhizobia. The significance and requirement of the very long chain fatty acids for outer membrane stability as well as the genetics of the synthesis pathway are discussed. The biological role of these fatty acids for bacterial life in extremely different environments (soil and intracellular space within nodules) is also considered.

Although the fungal ability to bind heavy metals has been known for many years, it was not determined which component is responsible for this process. The aim of the study was to isolate (1 → 3)-α- d -glucans from various fungi, select the most efficient compound in the biosorption of heavy metals, and determine its characteristics. The best α-glucan for treatment of aqueous solutions from heavy metals (Ni2+, Cd2+, Zn2+, Pb2+) is the polymer isolated from Shiitake, especially its SH 37 variety. Using various techniques, it was confirmed that the studied polysaccharide consists mainly of glucose molecules (75.9%), i.e. glucose α-anomers linked by (1 → 3)-linkage (86%). The well-developed surface, low crystallinity, and the large number of –OH groups provide this polymer with good sorption properties. Probably, the main mechanism of metal uptake is metabolism-independent process that depends on the content and spatial structure of the glucans present in their cell wall.

To obtain insight into physicochemical interactions between Cu(II) ions, kaolinite, and exopolysaccharide (EPS) synthesized by Sinorhizobium meliloti Rm 1021 soil bacteria, an adsorption, electrokinetic, and aggregation study was performed in the selected systems. The obtained data showed that supporting electrolyte type affects both EPS and Cu(II) ions adsorption. For initial Cu(II) concentration 100 mg/L, 4.36 ± 0.25 mg/g (21.80 ± 1.00%) of the ions were adsorbed in 0.001 M NaCl and 3.76 ± 0.20 mg/g (18.80 ± 1.00%) in 0.001 M CaCl2. The experimental data were best fitted to the Langmuir model as well as pseudo second-order equation. The EPS adsorbed amount on kaolinite was higher in the CaCl2 electrolyte than in NaCl one. For an initial polymer concentration of 100 mg/L, the EPS adsorbed amount was 4.69 ± 0.08 mg/g (23.45 ± 0.40%) in 0.001 M NaCl and 5.26 ± 0.15 mg/g (26.32 ± 0.75%) in 0.001 M CaCl2. In the mixed system, regardless of electrolyte type, exopolysaccharide contributed to immobilization of higher amount of copper(II) ions on the clay mineral. Also, in the samples containing heavy metal ions and exopolysaccharide simultaneously, the aggregation of kaolinite particles was the strongest. The results presented in the paper may be very helpful in soil bioremediation, especially in the development of technologies reducing the mobility of heavy metals in the environment.

This paper describes the electrokinetic, adsorption and stability properties of the silica–exopolysaccharide Sinorhizobium meliloti (EPS). Exopolysaccharide adsorbs on the silica surface in the whole pH range, but the adsorption amount increases with the pH growth (at pH 9 approx. 0.8 mg/m2). The EPS adsorption affects the electrokinetic and stability properties of the system. At low pH values (3 and 4.6) the increase in the system stability occurs (electrosteric interactions), whereas at higher (7.6 and 9) the light system destabilization was noted (formation of a single polymer bridges between particles).

Various phytochemical studies have revealed that jackfruit (Artocarpus heterophyllus Lam.) is rich in bioactive compounds, including carotenoids, flavonoids, volatile acids, tannins, and lectins. The aim of the study was to analyze the biological activity of water-soluble polysaccharide (WSP) isolated from jackfruit and to assess its immunomodulatory, cytotoxic, and anti-oxidative effects on human colon carcinoma cells in vitro. The neutral red (NR) uptake assay revealed no toxic influence of the polymer on the viability of tumor cells (HT29 and SW620). After 24 h and 48 h of incubation, the cellular viability was not lower than 94%. The metabolic activity of the cells (MTT) at the compound concentration of 250 &micro, g/mL was higher than 92% in comparison to the control. WSP (250 &micro, g/mL) exerted no significant effect on the morphology of the cells was determined by May-Gr&uuml, nwald-Giemsa staining. WSP changed nitric oxide (NOx) production by the tumor cells depending on the time of incubation and prior 2-h stimulation of the cells with E. coli 0111:B4 LPS. It significantly stimulated IL-1&beta, production by the tumor cells. The IL-6 level increased but that of IL-10 decreased by a WSP concentration-dependent manner. No such effect was detected in SW620. The WSP had antioxidant properties. In conclusion, water-soluble polysaccharide isolated from A. heterophyllus exhibits significant biological activity towards many types of both normal and cancerous cells. Therefore, it may be considered as a useful agent in the protection of human health or in functional and dietary nutrition.

A simple, low-cost, and reproducible method for creating materials with even silver nanoparticles (AgNP) dispersion was established. Chitosan nanofibers with silica phase (CS/silica) were synthesized by an electrospinning technique to obtain highly porous 3D nanofiber scaffolds. Silver nanoparticles in the form of a well-dispersed metallic phase were synthesized in an external preparation step and embedded in the CS/silica nanofibers by deposition for obtaining chitosan nanofibers with silica phase decorated by silver nanoparticles (Ag/CS/silica). The antibacterial activity of investigated materials was tested using Gram-positive and Gram-negative bacteria. The results were compared with the properties of the nanocomposite without silver nanoparticles and a colloidal solution of AgNP. The minimum inhibitory concentration (MIC) of obtained AgNP against Staphylococcus aureus (S. aureus) ATCC25923 and Escherichia coli (E. coli) ATCC25922 was determined. The physicochemical characterization of Ag/CS/silica nanofibers using various analytical techniques, as well as the applicability of these techniques in the characterization of this type of nanocomposite, is presented. The resulting Ag/CS/silica nanocomposites (Ag/CS/silica nanofibers) were characterized by small angle X-ray scattering (SAXS), X-ray diffraction (XRD), and atomic force microscopy (AFM). The morphology of the AgNP in solution, both initial and extracted from composite, the properties of composites, the size, and crystallinity of the nanoparticles, and the characteristics of the chitosan fibers were determined by electron microscopy (SEM and TEM).

Alkali-soluble polysaccharides (ASPs) were isolated from the cell wall of four Aspergillus species (A. fumigatus, A. nidulans, A. niger, and A. wentii). The chemical and spectroscopic investigations (immunofluorescent labelling, composition analysis, methylation analysis, FTIR, and 1H NMR) indicated that the ASPs were polymers composed almost exclusively of (1→3)-linked α-D-glucose. After carboxymethylation (CM), the activity of (1→3)-α-D-glucans on three human cell lines (HSF, HeLa, and Jurkat) was assessed. Anti-proliferative, cytotoxic, and free radical scavenging action of CM-α-D-glucans was analysed. All the tested CM-α-D-glucans decreased cellular metabolism. However, incubation with CM-α-D-glucan from A. wentii and A. niger increased (by ca. 50%) the viability of HSF cells. Moreover, an over 5-fold increase in the viability was found for Jurkat cells incubated with CM-α-D-glucans from A. fumigatus and A. nidulans. The CM-(1→3)-α-D-glucans from the tested Aspergillus species expressed no free radical scavenging action. Fluorescent staining revealed that CM-α-D-glucans exerted slight toxic effects on cell viability and no action on F-actin filaments of cellular cytoskeleton organization.

The chemical structures of the unusual hopanoid-containing lipid A samples of the lipopolysaccharides (LPS) from three strains of Bradyrhizobium (slow-growing rhizobia) have been established. They differed considerably from other Gram-negative bacteria in regards to the backbone structure, the number of ester-linked long chain hydroxylated fatty acids, as well as the presence of a tertiary residue that consisted of at least one molecule of carboxyl-bacteriohopanediol or its 2-methyl derivative. The structural details of this type of lipid A were established using one- and two-dimensional NMR spectroscopy, chemical composition analyses, and mass spectrometry techniques (electrospray ionization Fourier-transform ion cyclotron resonance mass spectrometry and MALDI-TOF-MS). In these lipid A samples the glucosamine disaccharide characteristic for enterobacterial lipid A was replaced by a 2,3-diamino-2,3-dideoxy-d-glucopyranosyl-(GlcpN3N) disaccharide, deprived of phosphate residues, and substituted by an α-d-Manp-(1→6)-α-d-Manp disaccharide substituting C-4' of the non-reducing (distal) GlcpN3N, and one residue of galacturonic acid (d-GalpA) α-(1→1)-linked to the reducing (proximal) amino sugar residue. Amide-linked 12:0(3-OH) and 14:0(3-OH) were identified. Some hydroxy groups of these fatty acids were further esterified by long (ω-1)-hydroxylated fatty acids comprising 26-34 carbon atoms. As confirmed by mass spectrometry techniques, these long chain fatty acids could form two or three acyloxyacyl residues. The triterpenoid derivatives were identified as 34-carboxyl-bacteriohopane-32,33-diol and 34-carboxyl-2β-methyl-bacteriohopane-32,33-diol and were covalently linked to the (ω-1)-hydroxy group of very long chain fatty acid in bradyrhizobial lipid A. Bradyrhizobium japonicum possessed lipid A species with two hopanoid residues.

Water-insoluble, alkali-soluble polysaccharide (marked as ASP) was extracted from the vegetative mycelium and fruiting body ofCerrena unicolorstrain. Monosaccharide examination of ASP demonstrated that the isolated biopolymer was composed mainly of glucose, xylose, and mannose monomers. The methylation investigation of studied polymers indicated that (1→3)-linkedα-D-Glcpis the major chain constituent (92.2% for glucans isolated from fruiting body and 90.1% from mycelium).1H NMR, FT-IR, and immunofluorescent labelling determinations confirmed that the polysaccharides isolated from both fruiting body and mycelium ofC. unicolorare (1→3)-α-d-glucans. The obtained (1→3)-α-d-glucans showed differences in viscosity and similar characteristics in optical rotations. (1→3)-α-d-Glucans extracted from mycelium and fruiting body ofC. unicolorwere also used as potential and specific inducers of mutanase synthesis byTrichoderma harzianum. The highest mutanase activity (0.38 U/mL) was obtained after induction of enzyme by (1→3)-α-d-glucan isolated from the mycelium ofC. unicolor, and this biopolymer has been suggested as a new alternative to streptococcal mutan for the mutanase induction inT. harzianum. (1→3)-α-d-Glucan-induced mutanase showed high hydrolysis potential in reaction with dextranase-pretreated mutan, where maximal degree of saccharification and solubilization of this bacterial homoglucan (83.1% and 78.4%, resp.) was reached in 3 h at 45°C.

Cellobiose dehydrogenase (CDH), an extracellular flavocytochrome produced by several wood-degrading fungi, was detected in the culture supernatant of the selective delignifier Pycnoporus sanguineus maintained on a cellulose-based liquid medium. Cellobiose dehydrogenase was purified as two active fractions: CDH1-FAD (flavin domain) (40.4 fold) with recovery of 10.9% and CDH1 (flavo-heme enzyme) (54.7 fold) with recovery of 9.8%. As determined by SDS-PAGE, the molecular mass of the purified enzyme was found to be 113.4kDa and its isoelectric point was 4.2, whereas these values for the FAD-domain were 82.7kDa and pI=6.7. The carbohydrate content of the purified enzymes was 9.2%. In this work, the cellobiose dehydrogenase gene cdh1 and its corresponding cDNA from fungus P. sanguineus were isolated, cloned, and characterized. The 2310bp full-length cDNA of cdh1 encoded a mature CDH protein containing 769 amino acids, which was preceded by a signal peptide of 19 amino acids. Moreover, both active fractions were characterized in terms of kinetics, temperature and pH optima, and antioxidant properties.

A water-soluble polysaccharide (WSP) extracted from mango (Mangifera indica L.) fruits has been suggested as a new alternative to mutan for mutanase induction in Trichoderma harzianum. Structural analyses revealed that the purified WSP was a (1→3),(1→4)-α-D-glucan with the molecular mass of ca. 760 kDa in which the (1→4)-linked and (1→3)-linked α-Glcp residues were in a ratio of 1:2.4. When the strain T harzianum CCM F-340 was grown in the presence of WSP, the maximal enzyme productivity obtained after 3 days of cultivation was 34 mU/mL. The mango WSP proved to be a very effective stimulus of mutanase expression giving a 5.1-fold higher than without WSP, transcription. It was shown that the mixture of WSP-induced mutanase and commercial dextranase had a high hydrolytic potential in the reaction with streptococcal mutan, where maximal degrees of solubilization and saccharification of this biopolymer (93.4% and 80%, respectively) were reached within 9h (solubilization) and 24h (saccharification). The mixed enzymatic preparation was also effective in degradation of streptococcal mutan and its removal from cariogenic biofilms. After 3h hydrolysis, only 18.2% of the biofilm remained adhered to the glass surface.

Chemical analyses and mass spectrometry were used to study the structure of the lipopolysaccharide (LPS) isolated from Aeromonas bestiarum strain K296, serotype O18. ESI-MS revealed that the most abundant A. bestiarum LPS glycoforms have a hexa-acylated or tetra-acylated lipid A with conserved architecture of the backbone, consisting of a 1,4′-bisphosphorylated β-(1→6)-linked d-GlcN disaccharide with an AraN residue as a non-stoichiometric substituent and a core oligosaccharide composed of Kdo1Hep6Hex1HexN1P1. 1D and 2D NMR spectroscopy revealed that the O-specific polysaccharide (OPS) of A. bestiarum K296 consists of a branched tetrasaccharide repeating unit containing two 6-deoxy-l-talose (6dTalp), one Manp and one GalpNAc residues; thus, it is similar to that of the OPS of A. hydrophila AH-3 (serotype O34) in both the sugar composition and the glycosylation pattern. Moreover, 3-substituted 6dTalp was 2-O-acetylated and additional O-acetyl groups were identified at O-2 and O-4 (or O-3) positions of the terminal 6dTalp. Western blots with polyclonal rabbit sera showed that serotypes O18 and O34 share some epitopes in the LPS. The very weak reaction of the anti-O34 serum with the O-deacylated LPS of A. bestiarum K296 might have been due to the different O-acetylation pattern of the terminal 6dTalp. The latter suggestion was further confirmed by NMR.

Oligotropha carboxidovorans strain OM5 (previously known as Pseudomonas carboxydovorans OM5) is a rod-shaped Gram-negative bacterium isolated from wastewater. This bacterium is able to live in aerobic and, facultatively, in autotrophic conditions. For autotrophic growth, the bacteria can utilize carbon monoxide or hydrogen as a source of energy. The O-specific polysaccharide isolated from O. carboxidovorans OM5 lipopolysaccharide was structurally characterized using chemical analyses, 1D and 2D NMR spectroscopy, and MALDI-TOF mass spectrometry techniques. The polysaccharide was found to be a homopolymer built up of 3- O -methyl-α- d -mannose residues linked by (1 → 2)-glycosidic bonds. The degree of polymerization of high-molecular-weight polysaccharide was estimated at approximately 35–40 units. The structure of the homopolymer is depicted below: [→2)-3- O Me-α- d -Man p -(1→] ∼35–40

Six rhizobium (Rhizobium leguminosarum bv. Trifolii TA1, Sinorhizobium meliloti 1021, Mesorhizobium huakuii IFO 15243T, Ochrobactrum lupini LUP 21T, Bradyrhizobium japonicum USDA110 and B. elkaniiUSDA 76) and two Escherichia coli strains (E. coli ATCC 25922 and E. coli HB 101) were compared in respect to polymyxin B and EDTA resistance, as well as bacterial outer membrane (OM) permeability to a fluorescent hydrophobic agent (N-phenyl-1-naphthylamine - NPN). TEM (Transmission Electron Microscopy) and a microbial test demonstrated that all the rhizobia were much more resistant to polymyxin B in comparison with E. coli strains. EDTA and polymyxin B enhance permeability of B. japonicum and O. lupini OM. Other rhizobia incorporated NPN independently of the presence of membrane-deteriorating agents; however, the level of fluorescence (measured as NPN absorption) was strain dependent.

This review focuses on the chemistry and structures of (Brady)rhizobium lipids A, indispensable parts of lipopolysaccharides. These lipids contain unusual (ω-1) hydroxylated very long chain fatty acids, which are synthesized by a very limited group of bacteria, besides rhizobia. The significance and requirement of the very long chain fatty acids for outer membrane stability as well as the genetics of the synthesis pathway are discussed. The biological role of these fatty acids for bacterial life in extremely different environments (soil and intracellular space within nodules) is also considered.

The O-polysaccharide was obtained from the lipopolysaccharide of the stem-nodulating nitrogen-fixing bacterium Azorhizobium caulinodans HAMBI 216 and studied by sugar and methylation analyses along with 1 H and 13 C NMR spectroscopy. The polysaccharide was found to have a linear pentasaccharide repeating unit containing d -rhamnose and its rarely occurring 2- O -methyl (Rha2 O Me) and 3- C -methyl (Rha3 C Me) derivatives and having the following structure: →3)-α- d -Rha p 2 O Me-(1→2)-β- d -Rha p 3 C Me-(1→3)-α- d -Rha p -(1→2)-β- d -Rha p 3 C Me-(1→3)-α- d -Rha p -(1→

The structure of the lipid A of the lipopolysaccharide (LPS) from Azorhizobium caulinodans, a symbiont of the tropical legume Sesbania rostrata, was investigated by chemical compositional analyses, mass spectrometry, as well as 1D and 2D NMR spectroscopy techniques. The lipid A backbone was composed of a β-(1→6)-linked 2,3-diamino-2,3-dideoxy- d -glucopyranose (GlcpN3N) disaccharide and α- d -glucuronic acid (GlcpA). Nuclear magnetic resonance spectroscopy revealed that the GlcpA was connected to the reducing end of the diaminosugar disaccharide via an α-(1→1) glycosidic bond. The lipid A was deprived of phosphate residues. ESI-MS analysis showed that the lipid A preparation was a mixture of molecules due to the occurrence of different acylation patterns. The GlcpN3N disaccharide backbone was N-acylated at the C-2, C-3, C-2′ and C-3′ positions with 3-OH-18:0, 3-OH-14:0, 3-OH-20:1 and 3-OH-14:0 fatty acids, respectively. Nonpolar fatty acids as well as 3-OH-18:0 were found to be ester-linked. They were attached to hydroxyl groups of primary 3-OH fatty acids giving three acyloxyacyl moieties. Thus, the complete lipid A from A. caulinodans comprised seven acyl residues. Part of the lipid A molecules was esterified by 3-methoxybutyric acid. Azorhizobium caulinodans did not incorporate ω-1 hydroxylated very long chain fatty acids (e.g., 27-OH-28:0) into the lipid A, which makes this variant of endotoxin unusual among rhizobial lipids A.

The O-specific polysaccharide obtained by mild-acid degradation of the lipopolysaccharide from Aeromonas veronii bv. sobria strain K49 was studied by sugar and methylation analyses along with (1)H and (13)C NMR spectroscopy. The sequence of the sugar residues was determined using (1)H,(1)H NOESY and (1)H,(13)C HMBC experiments. The O-specific polysaccharide was found to be a high molecular mass polysaccharide composed of repeating units of the structure: →2)-β-D-Quip3NAc-(1→3)-α-L-Rhap-(1→3)-α-L-Rhap-(1→2)-α-L-Rhap-(1→3)-α-D-FucpNAc-(1→ ESI MS confirmed the pentasaccharide structure of the repeating unit, as the molecular mass peaks seen in the spectrum differed by 812.34 u, a value corresponding to the calculated molecular mass of the O-unit.

Purpose of work To show biological activity of carboxymethylated α-(1 → 3)-d-glucans isolated from the selected macromycetes fungi on human tumor and normal cells. Water-insoluble, alkali-soluble polysaccharides (WIP) were isolated from fruiting bodies of four macromycetes fungi: Lentinus edodes, Pleurotus ostreatus, Piptoporus betulinus and Laetiporus sulphureus. The structure of the polysaccharides was determined using composition analysis, methylation analysis, fourier transform infrared spectroscopy, and nuclear magnetic resonance spectroscopy. The chemical and spectroscopic investigations indicated that the polysaccharides were an α-(1 → 3)-d-glucans. A biological activity analysis of the carboxymethylated (CM) α-(1 → 3)-d-glucans was based on an assessment of their cytotoxic, mitochondrial metabolism-modulating, and free radical scavenging effects. The cytotoxic activity of the CM-glucans was concentration- and cell-type-dependent. The tested CM-glucans, generally, did not have a free radical scavenging effect. The CM-α-(1 → 3)-d-glucans isolated from the selected macromycetes fungi are biologically active and may therefore be used as diet or therapy supplements.

The endotoxic activities of lipopolysaccharides (LPS) isolated from different strains of rhizobia and rhizobacteria (Bradyrhizobium, Mesorhizobium, and Azospirillum) were compared to those of Salmonella enterica sv. Typhimurium LPS. The biological activity of all the examined preparations, measured as Limulus lysate gelation, production of tumor necrosis factor (TNF), interleukin-1β (IL-1β), and interleukin-6 (IL-6), and nitrogen oxide (NO) induction in human myelomonocytic cells (line THP-1), was considerably lower than that of the reference enterobacterial endotoxin. Among the rhizobial lipopolysaccharides, the activities of Mesorhizobium huakuii and Azospirillum lipoferum LPSs were higher than those of the LPS preparations from five strains of Bradyrhizobium. The weak endotoxic activity of the examined preparations was correlated with differences in lipid A structure compared to Salmonella.

The structure of lipid A from Azospirillum lipoferum, a plant-growth-promoting rhizobacterium, was investigated. It was determined by chemical analysis, mass spectrometric methods, as well as 1D and 2D NMR spectroscopy. Because of the presence of substituents, the investigated lipid A differs from typical enterobacterial lipid A molecules. Its backbone is composed of a beta-(1,6)-linked D-glucosamine disaccharide but lacks phosphate residues. Moreover, the reducing end of the backbone (position C-1) is substituted with alpha-linked d-galacturonic acid. 3-hydroxypalmitoyl residues are exclusively connected to amino groups of the glucosamine disaccharide. Hydroxyls at positions C-3 and C-3' are esterified with 3-hydroxymyristic acids. Primary polar fatty acids are partially substituted by nonpolar fatty acids (namely, 18:0, 18:1 or 16:0), forming acyloxyacyl moieties.

The O-specific polysaccharide was obtained from the lipopolysaccharide of the legume-endosymbiotic bacterium Ochrobactrum cytisi strain ESC1T and studied by chemical analyses and 1D and 2D NMR spectroscopy. The polysaccharide was found to have a disaccharide repeating unit containing α- d -fucose and β-N-acetyl- d -galactosamine residues connected via (1→3)-glycosidic bonds, resulting in the following structure: →3)-α- d -Fucp-(1→3)-β- d -GalpNAc-(1→ The d -GalpNAc residue was nonstoichiometrically substituted with a 4-O-methyl group (∼10%) or with a 4,6-O-(1-carboxy)-ethylidene residue (pyruvyl group) (∼10%).

The O-specific polysaccharides of Phyllobacterium trifolii PETP02 T , a microsymbiont of Trifolium pratense , were obtained by mild acid hydrolysis of the lipopolysaccharide and studied by chemical analyses, mass spectrometry, and 1 H and 13 C NMR spectroscopy, including homonuclear ( 1 H, 1 H DQF-COSY, TOCSY, NOESY) and heteronuclear ( 1 H, 13 C HSQC, HMQC, HMBC) experiments. It was revealed that α- d -rhamnose and β-3- C -methyl- d -rhamnose (evalose) were the only components of two identified O-polysaccharides. The major O-polysaccharide was found to consist of linear hexasaccharide repeating units, while the other minor one, is composed of disaccharide repeats. The following structures of two O-polysaccharides were established: →2)-β- d -Rha p 3 C Me-(1→3)-α- d -Rha p -(1→3)-α- d -Rha p -(1→2)-β- d -Rha p 3 C Me-(1→3)-α- d -Rha p -(1→2)-α- d -Rha p -(1→ and →2)-β- d -Rha p 3 C Me-(1→3)-α- d -Rha p -(1→

The chemical structure of the free lipid A isolated from Mesorhizobium huakuii IFO 15243T was elucidated. Lipid A is a mixture of at least six species of molecules whose structures differ both in the phosphorylation of sugar backbone and in fatty acylation. The backbone consists of a β (1′6) linked 2,3-diamino-2,3-dideoxyglucose (DAG) disaccharide that is partly substituted by phosphate at position 4′. The aglycon of the DAG-disaccharide has been identified as α-d-galacturonic acid. All lipid A species carry four amide-linked 3-hydroxyl fatty residues. Two of them have short hydrocarbon chains (i.e. 3-OH-i-13:0) while the other two have longer ones (i.e. 3-OH-20:0). Distribution of 3-hydroxyl fatty acids between the reducing and nonreducing DAG is symmetrical. The nonpolar as well as (ω-1) hydroxyl long chain fatty acids are components of acyloxyacyl moieties. Two acyloxyacyl residues occur exclusively in the nonreducing moiety of the sugar backbone but their distribution has not been established yet. The distal DAG amide-bound fatty acid hydroxyls are not stoichiometrically substituted by ester-linked acyl components.

Fluoroquinolones, relatively new chemotherapeutics, are widely used in the treatment of both human and veterinary diseases because of their broad spectrum of antibacterial activity and good pharmacokinetic properties. Enrofloxacin is used in veterinary medicine, while its main metabolite, ciprofloxacin, is one of the most frequently used human antibiotics in the world. Hence, there is a need for assays capable of determining and distinguishing between these drugs. Thin‐layer chromatography–bioautography (TLC–B) combines TLC with microbiological detection. In the present paper, semiquantitative determination of enrofloxacin and ciprofloxacin standards by TLC–B using Chrom Biodip® Antibiotics Test Kit is presented. The optimal conditions for bioautographic detection were established. The exponential relations were proposed for approximation of dependencies between areas of zone inhibitions and logarithm of the amounts applied. The detection limit for standards of both enrofloxacin and ciprofloxacin...

Oligoglucose molecules isolated from Azorhizobium caulinodans were characterized by compositional analysis, Smith degradation, matrix-assisted laser desorption/ionization time of flight mass spectrometry, and (1)H and (13)C nuclear magnetic resonance analysis. A. caulinodans produced nonbranched and unsubstituted cyclic glucans composed solely of glucose, with the degree of polymerization ranging from 10 to 13. A major fraction of the periplasmic glucans contains 11 glucose residues within rings. The glucose residues are linked by beta-(1,3) and beta-(1,6) glycosidic bonds. These molecules seem to be quite similar to the periplasmic beta-(1,3);(1,6)-glucans synthesized by the Bradyrhizobium strain and are substantially different from the cyclic beta-(1,2)-glucans produced by Agrobacterium and Sinorhizobium species. Azorhizobial cyclic glucan synthesis is not osmoregulated. The response to the osmotic stress in Azorhizobium can be regulated similarly to Brucella spp. It is probable that the biosynthesis of beta-glucans is subject to the feedback control mechanism.

Thin-layer chromatography combined with bioautography (TLC-B) is a screening technique which combines TLC with microbiological detection and results in enhanced sensitivity. In the work discussed in this paper TLC-B was used to detect two veterinary antibiotics, doxycycline and flumequine, in milk. The sensitivity of bioautography was compared with that of UV detection. All procedures, i.e. clean-up, concentration, separation, and detection, were performed on the same TLC plate.

Polar lipid compositions of seven strains belonging to the four species of the Mesorhizobium genus were described. The lipid patterns of Mesorhizobium strains were very similar. Only quantitative differences were observed. Diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), and phosphatidylcholine (PC) were found to be the major phospholipids of the analysed bacteria. In addition, two methylated derivatives of PE were observed: phosphatidyl-N,N-dimethylethanolamine (DMPE) and phosphatidyl-N-monomethylethanolamine (MMPE). Polar head groups of those phospholipids were predominately acylated with lactobacillic (19:0 cyclopropane) acid. Ornithine-containing lipid (OL) was also identified. 3-hydroxy fatty acids found in the lipid preparations were derived exclusively from the ornithine lipid. 3-hydroxylactobacillic was the main acyl residue amide linked to the ornithine.

The impact of five carbohydrate sources (glucose, maltose, galactose, sucrose, and lactose) on the chemical composition, structure, morphology, and physicochemical properties, as well as, viscosity of exopolysaccharides (EPSs) produced by Lactobacillus rhamnosus E/N was investigated. GLC-MS analysis and 2DNMR spectroscopy showed that the EPSs had the same primary structure independently of the carbon source used in the growth medium. The following EPS composition was elucidated: four rhamnose, two glucose, and one galactose residue with a pyruvate substituent. Molecular masses (M(w)) were determined by gel permeation chromatography, which revealed differences in M(w) distribution. EPS-Gal, EPS-Suc, and EPS-Lac showed heterogenic fractions of a high and low molecular weight, while EPS-Mal and EPS-Glc contained only a high-molecular-weight fraction. AFM microscopy revealed morphological differences in chain length, thickness, and branching. Differences in the Mw ratio and thickness of the polymer chain were correlated with high viscosity of EPS solutions. Our results indicate that a single bacterial strain, depending on the carbon source in the medium, can produce EPSs of different rheological properties.

Chromium(III) oxide is an amphoteric, dark green solid. This most stable dye is widely used in construction and ceramic industries as well as in painting. In this study, the attempt is made to determine flocculating properties of exopolysaccharide (EPS) synthesized by the bacteria Sinorhizobium meliloti 1021, which would increase the efficiency of chromium(III) oxide removal from sewages and wastewaters. The conditions under which EPS is the most effective destabilizing component of chromium(III) oxide suspension have been determined too. In order to characterize the structure of electric double layer formed at the solid/supporting electrolyte (EPS) solution interface, electrokinetic potential measurements and potentiometric titration were performed. The EPS amount adsorbed on the chromium(III) oxide surface as a solution pH function was also measured. Moreover, the stability of Cr2O3 suspension in the absence and presence of S. meliloti 1021 EPS was estimated. The pooled analysis of all obtained results showed that EPS causes chromium(III) oxide suspension destabilization in the whole examined pH range. The largest change in the system stability before and after the polymer addition was observed at pH 9. It is probable that under these conditions bridging flocculation occurs in the examined system.