Evgeny Kudryashov

Complex formation between DNA and cationic surfactant

Complex formation between short DNA fragments and dodecyltrimethylammonium bromide (DoTAB) has been studied using dynamic (DLS), static light scattering (SLS), high performance capillary electrophoresis (HPCE) and a DoTAB-specific electrode. The combination of diffusion coefficient obtained from DLS, mobility from HPCE and binding isotherm allow us to deduce information about the structure of complexes as a function of bound surfactant. The binding of cationic surfactant to short DNA fragments was shown to proceed in two stages. In the first stage of binding, surfactant ions exchange with counterions “condensed” on the surface of DNA. At this stage the effective charge on DNA does not change as shown by the combination of mobility and diffusion coefficients of the complexes. The observed high cooperativity of binding can be explained by hydrophobic interactions between the surfactant molecules. In the second stage, surfactant molecules bind to DNA without exchange of condensed counterions, which brings about a dramatic change in the effective charge of DNA with phase separation occurring at high DNA concentration.

Ultrasonic high-resolution longitudinal and shear wave measurements in food colloids: monitoring of gelation processes and detection of pathogens

In the present paper we describe the applications of the ultrasonic high-resolution longitudinal and shear wave measurements for food and bio-colloids. In the first example, both ultrasonic methods were used for the monitoring of the acidified milk gelation induced by glucono-δ-lactone (formation of gel network in yoghurt). Ultrasonic measurements demonstrated a high sensitivity to pre-gelation and gelation processes during the formation of acid milk gels. The hydration of colloidal calcium phosphate released into serum and the swelling of casein in micelles at pH 5.6–5.0 are suggested as the main contributors to the ultrasonic velocity and attenuation changes during the pre-gelation. The increase in shear loss modulus of acidified milk at pH 5.0-4.85 can be explained by the aggregation of the casein micelles into clusters. Subsequent reformation of these clusters into a gel network at pH 4.85–4.6 is observed as a sharp rise in the storage moduli of acid milk gels and an increase in the ultrasonic velocity. The second example is the application of the ultrasonic shear wave measurements for the detection of Salmonella in liquids. The antigen-antibody binding monitored by impedance measurements of a quartz crystal at 5, 15 and 25 MHz results in both the decrease in resonant frequency and an increase in the imaginary part of the quartz impedance. The analysis of the data indicates that the bacteria cells on the sensor surface do not exhibit pure mass-load behaviour, and the viscoelastic properties of the interfacial layer must be taken into account for quantitative analysis. Overall, our ultrasonic measurements demonstrate their high potential as non-destructive methods of analysis of complex foods and bio-colloids.

High-frequency shear and volume viscoelastic moduli of casein particle gel

Abstract Protein particle gels, which consist of a continuous three-dimensional network of flocculated particles, such as casein micelles, are the main constituents of many biological and food colloids. These gels have an extremely short region of linear elastic behaviour and small fracture strain, thus creating complications for their rheological analysis. In the present paper we describe the application of low amplitude ultrasonic measurements for the monitoring of casein particle gel formation in a suspension of casein particles (a process similar to the formation of gel network in yoghurt). Combination of high-resolution longitudinal and shear wave measurements allowed us to make a complete analysis of the shear and volume viscoelasticity of casein particle gels in the frequency range 5–25 MHz. Both shear moduli, the storage ( G ′ Gel ∼30 kPa, at 7 MHz) and the loss moduli ( G ″ Gel ∼100 kPa, at 7 MHz) of the gel in the megahertz frequency range are several orders higher than those determined previously by dynamic rheology at low frequency (0.1 Hz), indicating different contributions of the gel network to the changes in the viscoelastic parameters at low and high frequencies. The volume storage ( K ′ Gel ∼75 kPa, at 7 MHz) and loss ( K ″ Gel ∼15 kPa, at 7 MHz) moduli of the casein particle gel are of the same order as the shear modili of the gel. The contribution of the gel to the volume storage modulus of the whole suspension of casein particles is very small, about 0.001%. This indicates the importance of high-resolution ultrasonic measurements for analysis of these systems.

Surfactant-DNA complexes in low ionic strength dilute solutions

We have used a combination of optical spectroscopy, fluorescence, isothermal titration calorimetry, surfactant-selective electrode techniques, high-resolution ultrasonic velocity and density measurements to study the binding of the cationic surfactant dodecyltrimethylammonium bromide to short fragments, 200 base pairs, of DNA in dilute solutions at 1 mM NaCl concentration. At the first stage the surfactant forms small aggregates on the DNA surface. This binding is accompanied by a significant change in the DNA secondary structure. At the second stage, we observed the formation of large aggregates of surfactant on the DNA surface.

Excited state complex formation between 3-aminophthalhydrazide and DNA: a fluorescence quenching reaction

Abstract Fluorescence quenching reaction of 3-aminophthalhyrdazide (luminol) due to the interaction with short DNA fragments has been studied employing steady-state and time correlated single photon counting techniques. The interaction of luminol with DNA in the excited state has also been analysed using Stern–Volmer (S–V) mechanism. A very weak ground state interaction is proposed to explain the higher values of the quenching rate constant and is consistent with positive curvatures in the S–V plots. A plausible explanation of the quenching mechanism has been discussed on the basis of hydrogen bonding interaction between luminol and DNA fragment. It is proposed that the phosphate backbone present in DNA is responsible for the interaction with luminol.

High-Resolution Ultrasonic Spectroscopy for Analysis of Industrial Emulsions and Suspensions

This article describes the application of high-resolution ultrasonic spectroscopy (HR-US) for the analysis of industrial emulsions and suspensions. The benefits of HR-US are discussed, including the ability to perform a direct analysis of emulsions and suspensions, which would otherwise be impossible, require significantly more effort in the laboratory, or produce erroneous results. The HR-US 102 spectrometer is also introduced. Manufacture of this laboratory-scale instrument is now possible due to recent technological advances in HR-US. The article outlines the principles of the HR-US technique and illustrates the application of the HR-US 102 spectrometer for analysis of absorption of ligands on the surface of particles, thermal stability, and effects of thermal history on microstructure of emulsions, crystallization, and particle sizing in diluted and concentrated emulsions.

Ultrasonic Monitoring of Enzyme Catalysis; Enzyme Activity in Formulations for Lactose-Intolerant Infants

The paper introduces ultrasonic technology for real-time, nondestructive, precision monitoring of enzyme-catalyzed reactions in solutions and in complex opaque media. The capabilities of the technology are examined in a comprehensive analysis of the effects of a variety of diverse factors on the performance of enzyme β-galactosidase in formulations for reduction of levels of lactose in infant milks. These formulations are added to infants milk bottles prior to feeding to overcome the frequently observed intolerance to lactose (a milk sugar), a serious issue in healthy development of infants. The results highlight important impediments in the development of these formulations and also illustrate the capability of the described ultrasonic tools in the assessment of the performance of enzymes in complex reaction media and in various environmental conditions.

High‐resolution longitudinal and shear wave spectroscopy for analysis of complex colloids

In the present paper we describe the applications of the high‐resolution ultrasonic longitudinal and shear wave resonator techniques for food and bio‐colloids. Our longitudinal wave technique allows us to measure the velocity and attenuation of ultrasonic waves with a resolution of 0.0001% for ultrasonic velocity and 0.1% for the ultrasonic attenuation in small volumes (0.1 to 1 ml) in the frequency range 4–20 MHz. The shear wave technique provides information on viscoelastic parameters of liquids and gels in the frequency range 4–25 MHz. Both techniques allow continuous, temperature ramp and automatic titration measurements for analysis of various processes in complex colloids. Examples of the application of both techniques for studying heat‐induced milk coagulation, phase transitions in milk fats, acid milk gelation, and the detection of food pathogens are described. Combination of both techniques shows a high sensitivity to various processes, e.g., pregelation and gelation, in food colloids and provide...