Alexey S. Vasilchenko

Facile preparation of amine and amino acid adducts of [60]fullerene using chlorofullerene C60Cl6 as a precursor.

We report a general synthetic approach to the preparation of highly functionalized amine and amino acid derivatives of [60]fullerene starting from readily available chlorofullerene C(60)Cl(6). The synthesized water-soluble amino acid derivative of C(60) demonstrated pronounced antiviral activity, while the cationic amine-based compound showed strong antibacterial action in vitro.

The activity of [60]fullerene derivatives bearing amine and carboxylic solubilizing groups against Escherichia coli : a comparative study

We report a comparative investigation of the antibacterial activity of two water-soluble fullerene derivatives bearing protonated amine (AF) and deprotonated carboxylic (CF) groups appended to the fullerene cage via organic linkers. The negatively charged fullerene derivative CF showed no tendency to bind to the bacterial cells and, consequently, no significant antibacterial activity. In contrast, the compound AF loaded with cationic groups showed strong and partially irreversible binding to the negatively charged Escherichia coli K12 TG1 cells and to human erythrocytes, also possessing negative zeta potential. Adsorption of AF on the bacterial surface was visualized by atomic force microscopy revealing the formation of specific clusters (AF aggregates) surrounding the bacterial cell. Incubation of E. coli K12 TG1 with AF led to a dose-dependent bactericidal effect with LD50 = 79.1 µM. The presence of human erythrocytes in the test medium decreased the AF antibacterial activity. Thus we reveal that the water-soluble cationic fullerene derivative AF possesses promising antibacterial activity, which might be utilized in the development of novel types of chemical disinfectants.

A zeta potential value determines the aggregate’s size of penta-substituted [60]fullerene derivatives in aqueous suspension whereas positive charge is required for toxicity against bacterial cells

BackgroundThe cause–effect relationships between physicochemical properties of amphiphilic [60]fullerene derivatives and their toxicity against bacterial cells have not yet been clarified. In this study, we report how the differences in the chemical structure of organic addends in 10 originally synthesized penta-substituted [60]fullerene derivatives modulate their zeta potential and aggregate’s size in salt-free and salt-added aqueous suspensions as well as how these physicochemical characteristics affect the bioenergetics of freshwater Escherichia coli and marine Photobacterium phosphoreum bacteria. Dynamic light scattering, laser Doppler micro-electrophoresis, agarose gel electrophoresis, atomic force microscopy, and bioluminescence inhibition assay were used to characterize the fullerene aggregation behavior in aqueous solution and their interaction with the bacterial cell surface, following zeta potential changes and toxic effects.ResultsDynamic light scattering results indicated the formation of self-assembled [60]fullerene aggregates in aqueous suspensions. The measurement of the zeta potential of the particles revealed that they have different surface charges. The relationship between these physicochemical characteristics was presented as an exponential regression that correctly described the dependence of the aggregate’s size of penta-substituted [60]fullerene derivatives in salt-free aqueous suspension from zeta potential value. The prevalence of DLVO-related effects was shown in salt-added aqueous suspension that decreased zeta potential values and affected the aggregation of [60]fullerene derivatives expressed differently for individual compounds. A bioluminescence inhibition assay demonstrated that the toxic effect of [60]fullerene derivatives against E. coli cells was strictly determined by their positive zeta potential charge value being weakened against P. phosphoreum cells in an aquatic system of high salinity. Atomic force microscopy data suggested that the activity of positively charged [60]fullerene derivatives against bacterial cells required their direct interaction. The following zeta potential inversion on the bacterial cells surface was observed as an early stage of toxicity mechanism that violates the membrane-associated energetic functions.ConclusionsThe novel data about interrelations between physicochemical parameters and toxic properties of amphiphilic [60]fullerene derivatives make possible predicting their behavior in aquatic environment and their activity against bacterial cells.

Investigation of Copper Nanoparticles Antibacterial Mechanisms Tested by Luminescent Escherichia coli Strains

The electrostatic interaction between positively charged copper nanoparticles aggregates (ζ = +15.9 ± 8.63 mV) and negatively charged surface of E. coli K12 TG1 cells (ζ = -50.0 ± 9.35 mV) has been established. The time-dependent decline of bacterial cells zeta potential and the coupled inhibition of constitutive bioluminescence level are the results of this interaction. The development of oxidative stress, probably defined by the electron transfer from the cytoplasmic membrane respiratory chains through membrane-integrated copper nanoparticle to the molecular oxygen, is shown as luminescence induction in superoxide- and peroxide-inducible E. coli K-12 MG1655 pSoxS::lux and pKatG::lux reporter strains. The final result of this process, which is responsible for the development of the bactericidal effect of copper nanoparticles, is DNA damage by active oxygen species detected by SOS-inducible E. coli pRecA::lux luminescent strain.

Humidity-Dependent Bacterial Cells Functional Morphometry Investigations Using Atomic Force Microscope

The effect of a relative humidity (RH) in a range of 93–65% on morphological and elastic properties of Bacillus cereus and Escherichia coli cells was evaluated using atomic force microscopy. It is shown that gradual dehumidification of bacteria environment has no significant effect on cell dimensional features and considerably decreases them only at 65% RH. The increasing of the bacteria cell wall roughness and elasticity occurs at the same time. Observed changes indicate that morphological properties of B. cereus are rather stable in wide range of relative humidity, whereas E. coli are more sensitive to drying, significantly increasing roughness and stiffness parameters at RH ≤ 84% RH. It is discussed the dependence of the response features on differences in cell wall structure of gram-positive and gram-negative bacterial cells.

An investigation into the interaction between carbon-based nanomaterials and Escherichia coli cells using atomic force microscopy

Using atomic force microscopy (AFM), the nature of the contact between a wide range of carbonbased nanomaterials (CBNs) and Escherichia coli cells is described and the consequences of such an interaction are estimated. It is shown that the contact of multiwall and a number of single-wall carbon nanotubes with the surface of a model microorganism carries a probabilistic nature and is not accompanied by changes in the morphology and viability of bacterial cells. The damage of the surface structures and the consequent destruction of Escherichia coli are observed upon contact with single-wall carbon nanotubes with low degrees of purification, which is presumably determined by the presence of technological impurities in CBNs. A significant enhancement of the affinity of C60 fullerenes functionalized by amine groups to the surface of model microorganisms, which also leads to the development of a bactericidal effect, is shown.

Atomic Force Microscopy Reveals a Morphological Differentiation of Chromobacterium violaceum Cells Associated with Biofilm Development and Directed by N-Hexanoyl-L-Homoserine Lactone

Chromobacterium violaceum abounds in soil and water ecosystems in tropical and subtropical regions and occasionally causes severe and often fatal human and animal infections. The quorum sensing (QS) system and biofilm formation are essential for C. violaceums adaptability and pathogenicity, however, their interrelation is still unknown. C. violaceums cell and biofilm morphology were examined by atomic force microscopy (AFM) in comparison with growth rates, QS-dependent violacein biosynthesis and biofilm biomass quantification. To evaluate QS regulation of these processes, the wild-type strain C. violaceum ATCC 31532 and its mini-Tn5 mutant C. violaceum NCTC 13274, cultivated with and without the QS autoinducer N-hexanoyl-L-homoserine lactone (C6-HSL), were used. We report for the first time the unusual morphological differentiation of C. violaceum cells, associated with biofilm development and directed by the QS autoinducer. AFM revealed numerous invaginations of the external cytoplasmic membrane of wild-type cells, which were repressed in the mutant strain and restored by exogenous C6-HSL. With increasing bacterial growth, polymer matrix extrusions formed in place of invaginations, whereas mutant cells were covered with a diffusely distributed extracellular substance. Thus, quorum sensing in C. violaceum involves a morphological differentiation that organises biofilm formation and leads to a highly differentiated matrix structure.

Novel haemoglobin-derived antimicrobial peptides from chicken (Gallus gallus) blood: purification, structural aspects and biological activity.

To purify and characterize antimicrobial peptides derived from the acid extract of Gallus gallus blood cells.

Atomic force microscopy recognition of protein A on Staphylococcus aureus cell surfaces by labelling with IgG–Au conjugates

Summary The labelling of functional molecules on the surface of bacterial cells is one way to recognize the bacteria. In this work, we have developed a method for the selective labelling of protein A on the cell surfaces of Staphylococcus aureus by using nanosized immunogold conjugates as cell-surface markers for atomic force microscopy (AFM). The use of 30-nm size Au nanoparticles conjugated with immunoglobulin G (IgG) allowed the visualization, localization and distribution of protein A–IgG complexes on the surface of S. aureus. The selectivity of the labelling method was confirmed in mixtures of S. aureus with Bacillus licheniformis cells, which differed by size and shape and had no IgG receptors on the surface. A preferential binding of the IgG–Au conjugates to S. aureus was obtained. Thus, this novel approach allows the identification of protein A and other IgG receptor-bearing bacteria, which is useful for AFM indication of pathogenic microorganisms in poly-component associations.

Toxicity of carbon-based nanomaterials against Escherichia coli depends on dispersion efficacy of their water suspensions

The relationships between surface wettability, dispersion efficacy in water suspensions and biotoxicity of nine carbon-based nanomaterials (CBN) samples represented by nanotubes, nanofibres and fullerenes are established. It is shown that presence of polar groups on the surface of similar in structure CBNs increases their hydrophilicity, reduces the particles size in water suspensions, and manifests itself by growth of the toxicity level in the luminescent screening assay based on Escherichia coli with the cloned luxCDABE-genes Photobacterium leiognathi. The artificial increase in individual CBN dispersion efficacy by primary suspension in the aprotic solvent dimethyl sulfoxide with the following transfer into an aqueous environment, also has led to growth of the registered biological activity. At the same time, the dispersion efficacy of morphologically diverse CBN is not the main cause of distinctions in their biotoxicity..