Effect of pH-Regulation on the Capture of Lipopolysaccharides from E. coli EH100 by Four-Antennary Oligoglycines in Aqueous Medium.

Bacterial lipopolysaccharides (LPS) are designated as endotoxins, because they cause fever and a wide range of pathologies in humans. It is important to develop effective methodologies to detect trace quantities of LPS in aqueous systems. The present study develops a fine-tuning procedure for the entrapment of trace quantities of LPS from E. coli EH100. The capture agents are self-assemblies (tectomers) formed by synthetic four-antennary oligoglycine (C-(CH2-NH-Gly7)4, T4). Based on previously performed investigations of bulk and adsorption-layer properties of aqueous solutions containing T4 and LPS, the optimal conditions for the entrapment interactions are further fine-tuned by the pH regulation of aqueous systems. A combined investigation protocol is developed, including dynamic light scattering, profile analysis tensiometry, microscopic thin-liquid-film techniques, and transmission electron microscopy. The key results are: (1) two types of complexes between T4 and LPS are generated-amphiphilic species and "sandwich-like" hydrophilic entities; the complexes are smaller at lower pH, and larger at higher pH; (2) an optimum range of pH values is established within which the whole quantity of the LPS is entrapped by the tectomers, namely pH = 5.04-6.30. The obtained data substantiate the notion that T4 may be used for an effective capture and the removal of traces of endotoxins in aqueous systems.

Electric polarizability changes during E. coli culture growth.

The electric polarizability of bacteria has two main components: surface-charge dependent (SChD) and Maxwell-Wagner (MW). It has been reported that the low frequency SChD component of Escherichia coli K12 still arise in the frequency range 20kHz - 2MHz, together with the high-frequency MW one. All the previous experiments were carried out with bacterial cultures of E. coli K12 in the stationary phase. In the present work we study electric polarizability during culture growth with the aim of finding out how it is influenced by the physiological state of the cells. The electro-optical method of electric turbidimetry is used, which is based on the change in the optical density as a result of orientation of bacterial cells under the action of an applied electric field. Our results show that until the cell concentration increases exponentially, the polarizability and the cell size change synchronously, so that the polarizability is approximately a quadratic function of the average bacterial length. We explain this with dominance of the SChD component. However, that after the polarizability deceases twofold at insignificant length oscillations and the power of the function decreases to 1.5. The last result is interpreted as an increase in the MW component.

Influence of the medium electrolyte concentration on the electric polarizability of bacteria Escherichia coli in presence of ethanol.

The electric polarizability is an important parameter of bacteria, giving information about the electric properties of the cells. In our previous works [A.M. Zhivkov, A.Y. Gyurova, Colloids Surf. B: Biointerfaces 66 (2008) 201; A.Y. Gyurova, A.M. Zhivkov, Biophys. Chem., 139 (2009) 8; A.M. Zhivkov, A.Y. Gyurova, J. Phys. Chem. B, 113 (2009) 8375] we have applied an experimental approach to distinguish the contribution of the components of the two types of interface electric polarizability-surface charge dependent (ChD) and Maxwell-Wagner (MW) polarizability. It is based on electro-optical study of the separate influence of the outer and inner medium electrolyte concentration, which changes the external ChD and internal MW components of polarizability; the last effect is reached by the membrane permeability increase in low ethanol concentration. In the present work we investigate the behavior of electric polarizability of Escherichia coli K12 at increasing the outer KCl concentration in presence of 10 vol.% ethanol in order to check if the polarizability components change independently from one another. The conclusion is that the outer electrolyte concentration influence indirectly the internal MW component by the trans-membrane concentration gradient, but the polarizability components themselves change independently.

Influence of cytoplasm electrolyte concentration on Maxwell-Wagner polarizability of bacteria E. coli.

The electric polarizability of bacteria is considered in the literature to have a surface charge dependent (ChD) and a Maxwell-Wagner (MW) mechanism. We distinguish experimentally both the types of interface polarizability by the frequency of the electric field and the medium electrolyte concentration. It was shown in a previous work ( Zhivkov , A. M. ; Gyurova , A. Y. Colloids Surf., B 2008 , 66 , 201. ) that the ChD component is shown up on the outer bacteria surface even at megahertz frequencies. The MW polarizability is studied in the present work in the range from 20 kHz to 20 MHz by change in the inner (cytoplasm) electrolyte concentration. The ion transport through the cytoplasmic membrane of alive and fixed by formaldehyde E. coli K12 is accelerated by adding of ethanol in low concentration. The frequency dependence and the kinetics of the electric polarizability and the size of the bacteria are investigated by conservative electric dichroism, based on the alteration of the optical density at orientation of the cells in electric field. The conclusion is that the internal MW component has the main contribution to the change in the total bacteria polarizability, as well as the external MW and the internal ChD components are not shown up.

Electric dipole moments of Escherichia coli HB 101.

The theoretical and experimental studies of the particles' electric dipole moments in the microscopic and submicroscopic size range show that in the case of polar and conductive media the interfacial components of the dipole moments are of greatest importance. While in the range of manometer's sizes there seems to be no important problems in the identification and in the estimation of the values of the dipole moments at present, in the micrometer range there are serious problems. In this communication these problems are considered and illustrated by electro-optic investigations of Escherichia coli HB 101.

Influence of ethanol on the high frequency electric polarizability of E. coli.

The interface electric polarizability of bacteria (charge dependent (ChD) and Maxwell-Wagner (MW) polarizabilities) gives information about their electric charge, determined by the structure and functional state. It is well known that the polarizability could be changed significantly by adding some substances to the suspension, and can be measured using an electro-optical (EO) method. There are some literature data, according to which the adding of ethanol decreases the electric polarizability of the cells. However the reason for the change in this parameter is not clear, as well as which component (ChD and/or MW) of polarizability has the main contribution. Generally the present work shows that the effect of ethanol is connected to the change of the internal (cytoplasm) MW polarizability and is mainly caused by increasing the cell membrane permeability. This results in an ionic flow through the membrane, which velocity and direction depends on the relative values of the inner (cytoplasm) and the outer medium ionic strength.