The assignment of 11 B and 1 H resonances in the post-reaction mixture from the dry synthesis of Li(BH3 NH2 BH2 NH2 BH3 ).

We report a detailed 1 H NMR and 11 B NMR study of as synthesised Li ( BH 3 NH 2 BH 2 NH 2 BH 3 ) obtained in a novel dry-synthesis method. A combination of 1D and 2D single- and triple-quantum techniques was used for the assignment of all observed signals. Minor side-products and reactants were detected in the product: NH 3 BH 3 , Li ( NH 2 BH 3 ) , Li ( BH 4 ) , and two yet unknown salts containing 7-membered chain anions: ( BH 3 NH 2 BH 2 NH 2 BH 2 NH 2 BH 3 ) - and ( BH ( NH 2 BH 3 ) 3 ) - . We believe the assignment provided within this study might be helpful when analysing the mixtures containing numerous ammonia borane derivatives, which often give overlapping signals that are hard to distinguish.

Regulatory and Enterotoxin Gene Expression and Enterotoxins Production in Staphylococcus aureus FRI913 Cultures Exposed to a Rotating Magnetic Field and trans-Anethole.

The study aimed to examine the influence of a rotating magnetic field (RMF) of two different frequencies (5 and 50 Hz) on the expression of regulatory (agrA, hld, rot) and staphylococcal enterotoxin (SE-sea, sec, sel) genes as well as the production of SEs (SEA, SEC, SEL) by the Staphylococcus aureus FRI913 strain cultured on a medium supplemented with a subinhibitory concentration of trans-anethole (TA). Furthermore, a theoretical model of interactions between the bacterial medium and bacterial cells exposed to RMF was proposed. Gene expression and SEs production were measured using quantitative real-time PCR and ELISA techniques, respectively. Based on the obtained results, it was found that there were no significant differences in the expression of regulatory and SE genes in bacteria simultaneously cultured on a medium supplemented with TA and exposed to RMF at the same time in comparison to the control (unexposed to TA and RMF). In contrast, when the bacteria were cultured on a medium supplemented with TA but were not exposed to RMF or when they were exposed to RMF of 50 Hz (but not to TA), a significant increase in agrA and sea transcripts as compared to the unexposed control was found. Moreover, the decreased level of sec transcripts in bacteria cultured without TA but exposed to RMF of 50 Hz was also revealed. In turn, a significant increase in SEA and decrease in SEC and SEL production was observed in bacteria cultured on a medium supplemented with TA and simultaneously exposed to RMFs. It can be concluded, that depending on SE and regulatory genes expression as well as production of SEs, the effect exerted by the RMF and TA may be positive (i.e., manifests as the increase in SEs and/or regulatory gene expression of SEs production) or negative (i.e., manifests as the reduction in both aforementioned features) or none.

The Assessment of Activity of Antiseptic Agents against Biofilm of Staphylococcus aureus Measured with the Use of Processed Microscopic Images.

Staphylococcal biofilms are major causative factors of non-healing wound infections. Their treatment algorithms recommend the use of locally applied antiseptic agents to counteract the spread of infection. The efficacy of antiseptics against biofilm is assessed in vitro by a set of standard quantitative and semi-quantitative methods. The development of software for image processing additionally allowed for the obtainment of quantitative data from microscopic images of biofilm dyed with propidium iodine and SYTO-9 reagents, differentiating dead cells from live ones. In this work, the method of assessment of the impact of antiseptic agents on staphylococcal biofilm in vitro, based on biofilms' processed images, was proposed and scrutinized with regard to clinically relevant antiseptics, polyhexanide, povidone-iodine and hypochlorite. The standard quantitative culturing method was applied to validate the obtained data from processed images. The results indicated significantly higher activity of polyhexanide and povidone-iodine than hypochlorite against staphylococcal biofilm. Taking into account the fact that in vitro results of the efficacy of antiseptic agents against staphylococcal biofilm are frequently applied to back up their use in hospitals and ambulatory units, our work should be considered an important tool; providing reliable, quantitative data in this regard.

The Effect of Rotating Magnetic Field on Susceptibility Profile of Methicillin-Resistant Staphylococcus aureus Strains Exposed to Activity of Different Groups of Antibiotics.

Methicillin-resistant strains of Staphylococcus aureus (MRSA) have become a global issue for healthcare systems due to their resistance to most ß-lactam antibiotics, frequently accompanied by resistance to other classes of antibiotics. In this work, we analyzed the impact of combined use of rotating magnetic field (RMF) with various classes of antibiotics (ß-lactams, glycopeptides, macrolides, lincosamides, aminoglycosides, tetracyclines, and fluoroquinolones) against nine S. aureus strains (eight methicillin-resistant and one methicillin-sensitive). The results indicated that the application of RMF combined with antibiotics interfering with cell walls (particularly with the ß-lactam antibiotics) translate into favorable changes in staphylococcal growth inhibition zones or in minimal inhibitory concentration values compared to the control settings, which were unexposed to RMF. As an example, the MIC value of cefoxitin was reduced in all MRSA strains by up to 42 times. Apart from the ß-lactams, the reduced MIC values were also found for erythromycin, clindamycin, and tetracycline (three strains), ciprofloxacin (one strain), gentamicin (six strains), and teicoplanin (seven strains). The results obtained with the use of in vitro biofilm model confirm that the disturbances caused by RMF in the bacterial cell walls increase the effectiveness of the antibiotics towards MRSA. Because the clinical demand for new therapeutic options effective against MRSA is undisputable, the outcomes and conclusions drawn from the present study may be considered an important road into the application of magnetic fields to fight infections caused by methicillin-resistant staphylococci.

Evaluation of Chemical Changes in Laboratory-Induced Colistin-Resistant Klebsiella pneumoniae.

This study evaluates the electrical potential and chemical alterations in laboratory-induced colistin-resistant Klebsiella pneumoniae, as compared to the susceptible strain using spectroscopic analyses. The minimal inhibitory concentration (MIC) of colistin, ζ-potential and chemical composition analysis of K. pneumoniae strains are determined. The results obtained for the K. pneumoniaeCol-R with induced high-level colistin resistance (MIC = 16.0 ± 0.0 mg/L) are compared with the K. pneumoniaeCol-S strain susceptible to colistin (MIC = 0.25 ± 0.0 mg/L). Fourier transform infrared (FTIR) and Raman spectroscopic studies revealed differences in bacterial cell wall structures and lipopolysaccharide (LPS) of K. pneumoniaeCol-R and K. pneumoniaeCol-S strains. In the beginning, we assumed that the obtained results could relate to a negative charge of the bacterial surface and different electrostatic interactions with cationic antibiotic molecules, reducing the affinity of colistin and leading to its lower penetration into K. pneumoniaeCol-R cell. However, no significant differences in the ζ-potential between the K. pneumoniaeCol-R and K. pneumoniaeCol-S strains are noticed. In conclusion, this mechanism is most probably associated with recognisable changes in the chemical composition of the K. pneumoniaeCol-R cell wall (especially in LPS) when compared to the susceptible strain.

Potato Juice, a Starch Industry Waste, as a Cost-Effective Medium for the Biosynthesis of Bacterial Cellulose.

In this work, we verified the possibility of valorizing a major waste product of the potato starch industry, potato tuber juice (PJ). We obtained a cost-effective, ecological-friendly microbiological medium that yielded bacterial cellulose (BC) with properties equivalent to those from conventional commercial Hestrin-Schramm medium. The BC yield from the PJ medium (>4 g/L) was comparable, despite the lack of any pre-treatment. Likewise, the macro- and microstructure, physicochemical parameters, and chemical composition showed no significant differences between PJ and control BC. Importantly, the BC obtained from PJ was not cytotoxic against fibroblast cell line L929 in vitro and did not contain any hard-to-remove impurities. The PJ-BC soaked with antiseptic exerted a similar antimicrobial effect against Staphylococcus aureus and Pseudomonas aeruginosa as to BC obtained in the conventional medium and supplemented with antiseptic. These are very important aspects from an application standpoint, particularly in biomedicine. Therefore, we conclude that using PJ for BC biosynthesis is a path toward significant valorization of an environmentally problematic waste product of the starch industry, but also toward a significant drop in BC production costs, enabling wider application of this biopolymer in biomedicine.

The Co-Culture of Staphylococcal Biofilm and Fibroblast Cell Line: The Correlation of Biological Phenomena with Metabolic NMR1 Footprint.

Staphylococcus aureus is one of the most prevalent pathogens associated with several types of biofilm-based infections, including infections of chronic wounds. Mature staphylococcal biofilm is extremely hard to eradicate from a wound and displays a high tendency to induce recurring infections. Therefore, in the present study, we aimed to investigate in vitro the interaction between S. aureus biofilm and fibroblast cells searching for metabolites that could be considered as potential biomarkers of critical colonization and infection. Utilizing advanced microscopy and microbiological methods to examine biofilm formation and the staphylococcal infection process, we were able to distinguish 4 phases of biofilm development. The analysis of staphylococcal biofilm influence on the viability of fibroblasts allowed us to pinpoint the moment of critical colonization-12 h post contamination. Based on the obtained model we performed a metabolomics analysis by 1H NMR spectroscopy to provide new insights into the pathophysiology of infection. We identified a set of metabolites related to the switch to anaerobic metabolism that was characteristic for staphylococcal biofilm co-cultured with fibroblast cells. The data presented in this study may be thus considered a noteworthy but preliminary step in the direction of developing a new, NMR-based tool for rapid diagnosing of infection in a chronic wound.

Rotating Magnetic Field Increases ß-Lactam Antibiotic Susceptibility of Methicillin-Resistant Staphylococcus aureus Strains.

Methicillin-resistant strains of Staphylococcus aureus (MRSA) have developed resistance to most ß-lactam antibiotics and have become a global health issue. In this work, we analyzed the impact of a rotating magnetic field (RMF) of well-defined and strictly controlled characteristics coupled with ß-lactam antibiotics against a total of 28 methicillin-resistant and sensitive S. aureus strains. The results indicate that the application of RMF combined with ß-lactam antibiotics correlated with favorable changes in growth inhibition zones or in minimal inhibitory concentrations of the antibiotics compared to controls unexposed to RMF. Fluorescence microscopy indicated a drop in the relative number of cells with intact cell walls after exposure to RMF. These findings were additionally supported by the use of SEM and TEM microscopy, which revealed morphological alterations of RMF-exposed cells manifested by change of shape, drop in cell wall density and cytoplasm condensation. The obtained results indicate that the originally limited impact of ß-lactam antibiotics in MRSA is boosted by the disturbances caused by RMF in the bacterial cell walls. Taking into account the high clinical need for new therapeutic options, effective against MRSA, the data presented in this study have high developmental potential and could serve as a basis for new treatment options for MRSA infections.

In Vitro Efficacy of Bacterial Cellulose Dressings Chemisorbed with Antiseptics against Biofilm Formed by Pathogens Isolated from Chronic Wounds.

Local administration of antiseptics is required to prevent and fight against biofilm-based infections of chronic wounds. One of the methods used for delivering antiseptics to infected wounds is the application of dressings chemisorbed with antimicrobials. Dressings made of bacterial cellulose (BC) display several features, making them suitable for such a purpose. This work aimed to compare the activity of commonly used antiseptic molecules: octenidine, polyhexanide, povidone-iodine, chlorhexidine, ethacridine lactate, and hypochlorous solutions and to evaluate their usefulness as active substances of BC dressings against 48 bacterial strains (8 species) and 6 yeast strains (1 species). A silver dressing was applied as a control material of proven antimicrobial activity. The methodology applied included the assessment of minimal inhibitory concentrations (MIC) and minimal biofilm eradication concentration (MBEC), the modified disc-diffusion method, and the modified antibiofilm dressing activity measurement (A.D.A.M.) method. While in 96-well plate-based methods (MIC and MBEC assessment), the highest antimicrobial activity was recorded for chlorhexidine, in the modified disc-diffusion method and in the modified A.D.A.M test, povidone-iodine performed the best. In an in vitro setting simulating chronic wound conditions, BC dressings chemisorbed with polyhexanide, octenidine, or povidone-iodine displayed a similar or even higher antibiofilm activity than the control dressing containing silver molecules. If translated into clinical conditions, the obtained results suggest high applicability of BC dressings chemisorbed with antiseptics to eradicate biofilm from chronic wounds.

Potential of Bacterial Cellulose Chemisorbed with Anti-Metabolites, 3-Bromopyruvate or Sertraline, to Fight against Helicobacter pylori Lawn Biofilm.

Helicobacter pylori is a bacterium known mainly of its ability to cause persistent inflammations of the human stomach, resulting in peptic ulcer diseases and gastric cancers. Continuous exposure of this bacterium to antibiotics has resulted in high detection of multidrug-resistant strains and difficulties in obtaining a therapeutic effect. The purpose of the present study was to determine the usability of bacterial cellulose (BC) chemisorbed with 3-bromopyruvate (3-BP) or sertraline (SER) to act against lawn H. pylori biofilms. The characterization of BC carriers was made using a N2 adsorption/desorption analysis, tensile strength test, and scanning electron microscopy (SEM) observations. Determination of an antimicrobial activity was performed using a modified disk-diffusion method and a self-designed method of testing antibacterial activity against biofilm microbial forms. In addition, bacterial morphology was checked by SEM. It was found that BC disks were characterized by a high cross-linking and shear/stretch resistance. Growth inhibition zones for BC disks chemisorbed with 2 mg of SER or 3-BP were equal to 26.5-27.5 mm and 27-30 mm, respectively. The viability of lawn biofilm H. pylori cells after a 4-h incubation with 2 mg SER or 3-BP chemisorbed on BC disks was ≥4 log lower, suggesting their antibacterial effect. SEM observations showed a number of morphostructural changes in H. pylori cells exposed to these substances. Concluding, SER and 3-BP chemisorbed on BC carriers presented a promising antibacterial activity against biofilm H. pylori cells in in vitro conditions.

Innate Immune Response against Staphylococcus aureus Preincubated with Subinhibitory Concentration of trans-Anethole.

The study aimed to analyze morphological and functional changes of Staphylococcus aureus cells due to trans-anethole (a terpenoid and the major constituent of fennel, anise, or star anise essential oils) exposition, and their consequences for human neutrophils phagocytic activity as well as IL-8 production (recognized as the major chemoattractant). The investigation included the evaluation of changes occurring in S. aureus cultures, i.e., staphyloxanthin production, antioxidant activities, cell size distribution, and cells composition as a result of incubation with trans-anethole. It was found that the presence of trans-anethole in the culture medium reduced the level of staphyloxanthin production, as well as decreased antioxidant activities. Furthermore, trans-anethole-treated cells were characterized by larger size and a tendency to diffuse in comparison to the non-treated cells. Several cell components, such as phospholipids and peptidoglycan, were found remarkably elevated in the cultures treated with trans-anethole. As a result of the aforementioned cellular changes, the bacteria were phagocytized by neutrophils more efficiently (ingestion and parameters associated with killing activity were at a higher level as compared to the control system). Additionally, IL-8 production was at a higher level for trans-anethole modified bacteria. Our results suggest that trans-anethole represents a promising measure in combating severe staphylococcal infections, which has an important translational potential for clinical applications.

Immobilization pattern of morphologically different microorganisms on bacterial cellulose membranes.

The aim of this study was to assess the immobilization pattern of microorganisms characterized by varying cell shapes and sizes (rod-shaped bacteria Lactobacillus delbruecki, spherical-shaped yeast Saccharomyces cerevisiae and hyphae forms of Yarrowia lipolytica) on bacterial cellulose of various material properties. The 'adsorption-incubation' method was used for the purposes of immobilization. The immobilization pattern included adsorption efficiency, ability of the immobilized cells to multiply within the carrier expressed as incubation efficiency and the degree of release of the immobilized cells from the carrier. The efficiency of adsorption and incubation was affected by the morphology of the immobilized cells and increased together with cellulose surface area. For smaller bacterial cells a higher level of loading was obtained on the same surface as compared to larger yeast cells. During incubation, the number of immobilized bacterial and yeast cells increased significantly in comparison to the number of cells adsorbed on the carrier during the adsorption step. Despite the morphological differences between the S. cerevisiae and Y. lipolytica cells, there were no statistically significant differences in the efficiency of adsorption and incubation. It was also revealed that the release ratio values obtained for L. delbruecki and S. cerevisiae increased along with cellulose surface area. Interestingly, Y. lipolytica cells in the pseudohyphae and hyphae forms penetrated deeply into the three-dimensional network of BC nanofibrils which prevented subsequent cell release. It was confirmed that carrier selection must be individually matched to the type of immobilized cells based especially on its porosity-related parameters.

Modification of Bacterial Cellulose with Quaternary Ammonium Compounds Based on Fatty Acids and Amino Acids and the Effect on Antimicrobial Activity.

In the present work, bacterial cellulose (BC) membranes have been modified with bioactive compounds based on long chain dimer of C18 linoleic acid, referred to as the dilinoleic acid (DLA) and tyrosine (Tyr), a natural amino acid capable of forming noncovalent cation-π interactions with positively charged ethylene diamine (EDA). This new compound, [EDA][DLA-Tyr], has been synthesized by simple coupling reaction, and its chemical structure was characterized by 1H NMR and Fourier transform infrared spectroscopy. The antimicrobial activity of a new compound against S. aureus and S. epidermidis, two cocci associated with skin and wound infections, was assessed. The [EDA][DLA-Tyr] impregnated BC exhibited strong and long-term antimicrobial activity against both staphylococcal species. The results showed a 57-66% and 56-60% reduction in S. aureus and S. epidermidis viability, respectively, depending on [EDA][DLA-Tyr] concentration used. Importantly, [EDA][DLA-Tyr] molecules were released gradually from the BC pellicle, while a reference antibiotic, erythromycine (ER), did not show any antibacterial activity against S. aureus and S. epidermidis after 48 h of soaking in deionized water. Thus, a combination of [EDA][DLA-Tyr] and BC could be a promising new class of wound dressing displaying both biocompatibility and antimicrobial activity.

Increased water content in bacterial cellulose synthesized under rotating magnetic fields.

The current study describes properties of bacterial cellulose (BC) obtained from Komagataeibacter xylinus cultures exposed to the rotating magnetic field (RMF) of 50 Hz frequency and magnetic induction of 34 mT for controlled time during 6 days of cultivation. The experiments were carried out in the customized RMF exposure system adapted for biological studies. The obtained BC displayed an altered micro-structure, degree of porosity, and water-related parameters in comparison to the non-treated, control BC samples. The observed effects were correlated to the duration and the time of magnetic exposure during K. xylinus cultivation. The most preferred properties in terms of water-related properties were found for BC obtained in the setting, where RMF generator was switched off for the first 72 h of cultivation and switched on for the next 72 h. The described method of BC synthesis may be of special interest for the production of absorbent, antimicrobial-soaked dressings and carrier supports for the immobilization of microorganisms and proteins.

Complete Series of Alkali-Metal M(BH3NH2BH2NH2BH3) Hydrogen-Storage Salts Accessed via Metathesis in Organic Solvents.

We report a new efficient way of synthesizing high-purity hydrogen-rich M(BH3NH2BH2NH2BH3) salts (M = Li, Na, K, Rb, Cs). The solvent-mediated metathetic synthesis applied here uses precursors containing bulky organic cations and weakly coordinating anions. The applicability of this method permits the entire series of alkali-metal M(BH3NH2BH2NH2BH3) salts (M = Li, Na, K, Rb, Cs) to be obtained, thus enabling their comparative analysis in terms of crystal structures and hydrogen-storage properties. A novel polymorphic form of Verkade's base (C18H39N4PH)(BH3NH2BH2NH2BH3) precursor was also characterized structurally. For all compounds, we present a comprehensive structural, spectroscopic, and thermogravimetric characterization (PXRD, NMR, FTIR, Raman, and TGA/DSC/MS).

Facile formation of thermodynamically unstable novel borohydride materials by a wet chemistry route.

A novel wet synthetic method utilizing weakly coordinating anions that yields LiCl-free Zn-based materials for hydrogen storage has recently been reported. Here we show that this method may also be applied for the synthesis of the pure yttrium derivatives, M[Y(BH4)4] (M = K, Rb, Cs). Moreover, it can be extended to the preparation of previously unknown thermodynamically unstable derivatives, Li[Y(BH4)4] and Na[Y(BH4)4]. Importantly, these two H-rich phases cannot be accessed by standard dry (mechanochemical) or solid/gas synthetic methods due to the thermodynamic obstacles. Here we describe their crystal structures and selected important physicochemical properties.

Effects of rotating magnetic field exposure on the functional parameters of different species of bacteria.

The aim of the present study was to determine the effect of the rotating magnetic field (RMF) on the growth, cell metabolic activity and biofilm formation by S. aureus, E. coli, A. baumannii, P. aeruginosa, S. marcescens, S. mutans, C. sakazakii, K. oxytoca and S. xylosus. Bacteria were exposed to the RMF (RMF magnetic induction B = 25-34 mT, RMF frequency f = 5-50 Hz, time of exposure t = 60 min, temperature of incubation 37 °C). The persistence of the effect of exposure (B = 34 mT, f = 50 Hz, t = 60 min) on bacteria after further incubation (t = 300 min) was also studied. The work showed that exposure to RMF stimulated the investigated parameters of S. aureus, E. coli, S. marcescens, S. mutans, C. sakazakii, K. oxytoca and S. xylosus, however inhibited cell metabolic activity and biofilm formation by A. baumannii and P. aeruginosa. The results obtained in this study proved, that the RMF, depending on its magnetic induction and frequency can modulate functional parameters of different species of bacteria.

Synthesis and antibacterial activity of Schiff bases and amines derived from alkyl 2-(2-formyl-4-nitrophenoxy)alkanoates.

A series of novel Schiff bases and secondary amines were obtained in good yields, as a result of the reductive amination of alkyl 2-(2-formyl-4-nitrophenoxy)alkanoates with both aniline and 4-methoxyaniline under established mild reaction conditions. Sodium triacetoxyborohydride as well as hydrogen in the presence of palladium on carbon were used as efficient reducing agents of the Schiff bases, in both direct and stepwise reductive amination processes. The Schiff bases, amines, and amine hydrochlorides were designed as potential antibacterial agents, and structure-activity relationship could be established following in vitro assays against Gram-positive and Gram-negative bacteria. The minimal inhibitory concentration and zone of inhibition were also determined. In these tests, some of Schiff bases and secondary amine hydrochlorides showed moderate-to-good activity against Gram-positive bacteria, including S. aureus, M. luteus, and S. mutans.

Hydrogen storage materials: room-temperature wet-chemistry approach toward mixed-metal borohydrides.

The poor kinetics of hydrogen evolution and the irreversibility of the hydrogen discharge hamper the use of transition metal borohydrides as hydrogen storage materials, and the drawbacks of current synthetic methods obstruct the exploration of these systems. A wet-chemistry approach, which is based on solvent-mediated metathesis reactions of precursors containing bulky organic cations and weakly coordinating anions, leads to mixed-metal borohydrides that contain only a small amount of "dead mass". The applicability of this method is exemplified by Li[Zn2(BH4)5] and M[Zn(BH4)3] salts (M=Na, K), and its extension to other systems is discussed.

Effects of 50 Hz rotating magnetic field on the viability of Escherichia coli and Staphylococcus aureus.

This study presents results of research on the influence of rotating magnetic field (RMF) of the induction of 30 mT and the frequency of 50 Hz on the growth dynamics and cell metabolic activity of E. coli and S. aureus, depending on the exposure time. The studies showed that the RMF caused an increase in the growth and cell metabolic activity of all the analyzed bacterial strains, especially in the time interval t = 30 to 150 min. However, it was also found that the optical density and cell metabolic activity after exposition to RMF were significantly higher in S. aureus cultures. In turn, the study of growth dynamics, revealed a rapid and a significant decrease in these values from t = 90 min) in the case of E. coli samples. The obtained results prove that RMF (B = 30 mT, f = 50 Hz) has a stimulatory effect on the growth and metabolic activity of E. coli and S. aureus. Furthermore, taking into account the time of exposure, stronger influence of RMF on the viability was observed in S. aureus cultures, which may indicate that this effect depends on the shape of the exposed cells.