5-Deazaalloxazine as photosensitizer of singlet oxygen and potential redox-sensitive agent.

Flavins are a unique class of compounds that combine the features of singlet oxygen generators and redox-dependent fluorophores. From a broad family of flavin derivatives, deazaalloxazines are significantly underdeveloped from the point of view of photophysical properties. Herein, we report photophysics of 5-deazaalloxazine (1a) in water, acetonitrile, and some other solvents. In particular, triplet excited states of 1a in water and in acetonitrile were investigated using ultraviolet-visible (UV-Vis) transient absorption spectroscopy. The measured triplet lifetimes for 1a were all on the microsecond time scale (≈ 60 µs) in deoxygenated solutions. The quantum yield of S1 → T1 intersystem crossing for 1a in water was 0.43 based on T1 energy transfer from 1a to indicaxanthin (5) acting as acceptor and on comparative actinometric measurements using benzophenone (6). 1a was an efficient photosensitizer for singlet oxygen in aerated solutions, with quantum yields of singlet oxygen in methanol of about 0.76, compared to acetonitrile ~ 0.74, dichloromethane ~ 0.64 and 1,2-dichloroethane ~ 0.54. Significantly lower singlet oxygen quantum yields were obtained in water and deuterated water (Ð¤Δ ~ 0.42 and 0.44, respectively). Human red blood cells (RBC) were used as a cell model to study the antioxidant capacity in vitro and cytotoxic activity of 1a. Fluorescence-lifetime imaging microscopy (FLIM) data were analyzed by fluorescence lifetime parameters and distribution for different parts of the emission spectrum. Comparison of multidimensional fluorescent properties of RBC under physiological-like and oxidative-stress conditions in the presence and absence of 1a suggests its dual activity as probe and singlet-oxygen generator and opens up a pathway for using FLIM to analyze complex intracellular behavior of flavin-like compounds. These new data on structure-property relationship contribute to the body of information required for a rational design of flavin-based tools for future biological and biochemical applications.

Activation of enzymatic ethanol oxidation by ATP hydrolysis energy transferred to ADH1A enzyme molecule over Müller cell intermediate filaments.

Presently exciton activation of enzymatic oxidation of ethanol by human alcohol dehydrogenase (ADH) 1A enzyme is reported. The ADH1A enzyme was activated by infrared (IR) excitons transferred over Müller cell (MC) intermediate filaments (IFs). These IR excitons were generated by energy liberated upon enzymatic ATP hydrolysis and transferred to IFs. Also, the emission spectrum was recorded of the electronically excited ADH1A…NAD+…EtOH complexes obtained by energy transfer from IR excitons that traveled along IFs. These results support the hypothesis that ATP hydrolysis energy may be transmitted in vivo in the form of IR excitons, over the network of IFs, both within and between cells.

Swelling and membrane potential dynamics of glial Müller cells.

Presently a detailed biophysical model describing reversible and irreversible swelling dynamics of Müller cells (MC) is reported. The model includes a biophysical block of ionic and neutral species transport via MC membrane, water transport induced by osmotic pressure and pressure generated by membrane deformations, MC membrane potential and membrane mechanical properties. The model describes reversible and irreversible MC swelling (MCS) using the same set of parameters. The model was used in fitting available experimental data, and produced numerical values of previously unknown model parameters, including those describing mechanical properties of Müller cell membrane (MCM) with respect to bending and stretching. Numerical experiments simulating MC swelling showed complex oscillation dynamics of the relevant parameters in physiological initial conditions. In particular, MC membrane potential (ΔΨMC) demonstrated complex oscillation dynamics, which may be described by a superposition of several oscillations with their periods in the milliseconds, 100-ms and seconds time ranges. Dynamics of reversible and irreversible MCS, and the transition criteria from reversible to irreversible MCS modes were determined in model simulations.

Theoretical approaches used in the modelling of reversible and irreversible mitochondrial swelling in vitro.

Existing theoretical approaches were considered that allow modelling of mitochondrial swelling (MS) dynamics. Simple phenomenological kinetic models were reviewed. Simple and extended biophysical and bioenergetic models that ignore mechanical properties of inner mitochondrial membrane (IMM), and similar models that include these mechanical properties were also reviewed. Limitations of these models we considered, as regards correct modelling of MS dynamics. It was found that simple phenomenological kinetic models have significant limitations, due to dependence of the kinetic parameter values estimated by fitting of the experimental data on the experimental conditions. Additionally, such simple models provide no understanding of the detailed mechanisms behind the MS dynamics, nor of the dynamics of various system parameters during MS. Thus, biophysical and bioenergetic models ignoring IMM mechanical properties can't be used to model the transition between reversible and irreversible MS. However, simple and extended biophysical models that include IMM mechanical properties allow modelling the transition to irreversible swelling. These latter models are still limited due to significantly simplified description of biochemistry, compared to those of bioenergetic models. Finally, a strategy of model development is proposed, towards correct interpretation of the mitochondrial life cycle, including the effects of MS dynamics.

Theoretical analysis of reversible and irreversible mitochondrial swelling invivo.

Theoretical biophysical model is reported for mitochondrial swelling (MS) dynamics in vivo. This newly developed model is based on the detailed biophysical model of MS dynamics in vitro, where mechanical properties of the inner mitochondrial membrane (IMM) were taken into account. The present model of MS dynamics in vivo is capable of analyzing MS dynamic transition from the reversible (physiological) to the irreversible (pathological) mode. This model was used to describe myocytes, assuming 1000 mitochondria distributed homogeneously over the sarcoplasm. Solute transport through the myocyte membrane was described by simplified phenomenological mechanisms of solute uptake and release. Biophysical processes occurring in mitochondria within cells were similar to those included in the earlier reported in vitro biophysical model of MS dynamics. Additionally, in vivo MS dynamics was simulated in different initial conditions, with results different from those of the in vitro model. Note that the presently reported model is the first attempt to develop a detailed biophysical model for the analysis of MS dynamics in vivo, capable of reproducing the transition between reversible and irreversible MS dynamics.

Temperature dependence of IR exciton emission spectra in Müller cell intermediate filaments.

Temperature dependences of IR exciton properties in Müller cell (MC) intermediate filaments (IFs) isolated from porcine retina were studied. It was found that the widths of the spectral emission bands in the 2500 cm-1 and 5000 cm-1 energy ranges grow with temperature. It was found that temperature effects on the bandwidth may be described by thermal activation of the low-frequency vibrational modes of the IFs. The average activation energies for the two IR bands were estimated. Considering the dynamics of IR emission, its buildup time was independent on the sample temperature, while its decay time decreased with temperature. Thus, the emission decay rate increased exponentially with the sample temperature. The mechanisms explaining the observed temperature effects were proposed and discussed. Taking into account that MC IFs are capable of transmitting ATP hydrolysis energy within and between cells, with these properties being apparently common for all IFs, these IFs may be used by cells for physical energy transport and communications. As presently reported, temperature effects upon IR exciton spectra should not affect these proposed physiological functions to any significant extent. Therefore, the currently reported data are important for improving our understanding of the physical communication mechanisms operating within and between cells.

Photo-activation of mitochondrial ATP synthesis.

ATP production by mitochondria isolated from Saccharomyces cerevisiae cells was accelerated upon both direct and indirect mitochondrial photo-activation (MPA). The extent of direct MPA was dependent on the wavelength of excitation light. Direct MPA was created by light in cytochrome c spectral absorption bands (440, 520 and 550 nm), this light was absorbed producing electronically excited cytochrome c, and the excitation energy of the latter was used in the ATP production chain. The activity of cytochrome c was tested with 600 nm light, where cytochrome c does not absorb, and thus ATP production rate remained the same as in darkness. Note that ATP production rates were significantly larger under light at 550, 520 and 440 nm. Therefore, photo-activation of cytochrome c was the first step of MPA synthesis of ATP. Indirect MPA of ATP production also proceeded via electronically excited cytochrome c, by energy transfer from electronically excited Co/BN film to cytochrome c located in the inner mitochondrial membrane (IMM). Co/BN excitons were generated by photons absorbed by the Co/BN film, which was not in contact with the mitochondrial sample. Next, these excitons propagated along the Co/BN film to the part of the film that was in contact with the mitochondrial sample. There the exciton energy was transferred to cytochrome c located in the IMM, producing electronically excited cytochrome c. Thus, excited cytochrome c was generated in a way different from that of direct MPA. Next, the energy of excited cytochrome c was used in activated ATP synthesis, with virtually the same effect for 519 and 427 nm excitation. Thus, the first step of ATP synthesis in indirect MPA was the exciton energy transfer from Co/BN film to cytochrome c located in the IMM, producing an electronically excited cytochrome c molecule. A phenomenological mechanism of direct and indirect MPA was proposed, and the model parameters were obtained by fitting the model to the experimental data. However, more information is needed before the detailed mechanism of ATP synthesis activation by electronically excited cytochrome c could be understood. The present results support the earlier proposed hypothesis of indirect MPA of ATP production in vertebrate retina in daylight.

Photophysical properties of alloxazine derivatives with extended aromaticity - Potential redox-sensitive fluorescent probe.

The spectral and photophysical properties of two four-ring alloxazine derivatives, naphtho[2,3-g]pteridine-2,4(1H,3H)-dione (1a) and 1,3-dimethylnaphtho[2,3-g]pteridine-2,4(1H,3H)-dione, (1b) were studied. The propensity of 1a for excited-state proton transfer reactions in the presence of acetic acid as a catalyst was also studied, showing no signature of the reaction occurring. In addition, quenching of 1a fluorescence by acetic acid was investigated. Singlet and triplet states and spectral data for 1a and 1b were calculated using density functional theory TD-DFT at B3LYP/6-31G(d) and UB3LYP levels. Finally, fluorescence lifetime imaging microscopy (FLIM) using 1a and 1b as fluorescence probes was applied to in vitro human red blood cells (RBCs) with and without tert-butyl hydroperoxide (TB) as an oxidising agent. To evaluate and compare the effects of 1a and 1b on the redox properties of RBCs, the fluorescence lifetime, amplitude and fractional intensities were calculated, and phasor plot analysis was performed. The results obtained show the appearance of a new proximal cluster in the phasor fingerprint of RBCs in the presence of 1b and a shorter fluorescence lifetime of RBCs in the presence of 1a.

Effects of pulsed electric fields on exciton propagation efficiency along Müller cell intermediate filaments. Possible separation mechanism of high- and low-contrast images by the eye-brain system.

In the current study, we tested a possible mechanism of low- and high-contrast image component discrimination by the vertebrate eye-brain system. Apparently the eye-brain system has to discriminate between the low-contrast image component formed by light scattered within the retina, due to interaction of photons with cells and their parts, and the high-contrast image component transmitted by excitons via the quantum mechanism. Presently, effects of pulsed electric fields applied to Müller cell (MC) intermediate filaments (IFs) on the efficiency of exciton propagation were explored. The effects of both pulse duration and amplitude were recorded. These experimental results show that the eye-brain system may be using signal modulation to discriminate between high- and low-contrast image components, improving our understanding of high-contrast vision in vertebrates.

Contrary to consensus, oxidation of ethanol by human alcohol dehydrogenase (ADH) 1A is activated by ATP.

Presently we report that enzymatic oxidation of ethanol (EtOH) by ADH1A alcohol dehydrogenase is strongly accelerated in presence of adenosine triphosphate (ATP), by up to the factor of 20 in vitro. This result provides a different look on the role of ATP in functioning of alcohol dehydrogenases (ADH), which until presently were a textbook example of enzymes not requiring ATP and successfully operating without it. However, ATP is available in every living cell and will activate reactions conducted by ADH enzymes in vivo. Therefore, the body of published literature describing properties of numerous ADH enzymes requires a thorough revision.

Reversible and irreversible mitochondrial swelling: Effects of variable mitochondrial activity.

An extended biophysical model was obtained by upgrading the previously reported one (Khmelinskii and Makarov, 2021). The upgraded model accommodates variations of solute transport rates through the inner mitochondrial membrane (IMM) within the mitochondrial population, described by a Gaussian distribution. However, the model may be used for any functional form of the distribution. The dynamics of system parameters as predicted by the current model differed from that predicted by the previous model in the same initial conditions (Khmelinskii and Makarov, 2021). The amount of change varied from one parameter to the other, remaining in the 1-38% range. The upgraded model fitted the available experimental data with a better accuracy (R = 0.993) compared to the previous model (R = 0.978) using the same experimental data (Khmelinskii and Makarov, 2021). The fitting procedure also estimated the Gaussian distribution parameters. The new model requires much larger computational resources, but given its higher accuracy, it may be used for better analysis of experimental data and for better prediction of MS dynamics in different initial conditions. Note that activities of individual mitochondria in mitochondrial populations should vary within biological tissues. Thus, the currently upgraded model is a better tool for biological and bio-medical applications. We believe that this model is much better adapted to the analysis of MS dynamics in vivo.

Reversible and irreversible mitochondrial swelling in vitro.

Mitochondrial activity as regards ATP production strongly depends on mitochondrial swelling (MS) mode. Therefore, this work analyzes reversible and irreversible MS using a detailed biophysical model. The reported model includes mechanical properties of the inner mitochondrial membrane (IMM). The model describes MS dynamics for spherically symmetric, axisymmetric ellipsoidal and general ellipsoidal mitochondria. Mechanical stretching properties of the IMM were described by a second-rank rigidity tensor. The tensor components were estimated by fitting to the earlier reported results of in vitro experiments. The IMM rigidity constant of ca. 0.008 dyn/nm was obtained for linear deformations. The model also included membrane bending effects, which were small compared to those of membrane stretching. The model was also tested by simulation of the earlier reported experimental data and of the system dynamics at different initial conditions, predicting the system behavior. The transition criteria from reversible to irreversible swelling were determined and tested. The presently developed model is applicable directly to the analysis of in vitro experimental data, while additional improvements are necessary before it could be used to describe mitochondrial swelling in vivo. The reported theoretical model also provides an idea of physically consistent mechanism for the permeability transport pore (PTP) opening, which depends on the IMM stretching stress. In the current study, this idea is discussed briefly, but a detailed theoretical analysis of these ideas will be performed later. The currently developed model provides new understanding of the detailed MS mechanism and of the conditions for the transition between reversible and irreversible MS modes. On the other hand, the current model provides useful mathematical tools, that may be successfully used in mitochondrial biophysics research, and also in other applications, predicting the behavior of mitochondria in different conditions of the surrounding media in vitro or cellular cyto(sarco)plasm in vivo. These mathematical tools are based on real biophysical processes occurring in mitochondria. Thus, we note a significant progress in the theoretical approach, which may be used in real biological systems, compared to the earlier reported models. Significance of this study derives from inclusion of IMM mechanical properties, which directly impact the reversible and irreversible mitochondrial swelling dynamics. Reversible swelling corresponds to reversible IMM deformations, while irreversible swelling corresponds to irreversible deformations, with eventual membrane disruption. The IMM mechanical properties are directly dependent on the membrane biochemical composition and structure. The IMM deformationas are induced by osmotic pressure created by the ionic/neutral solute imbalance between the mitochondrial matrix media and the bulk solution in vitro, or cyto(sarco)plasm in vivo. The novelty of the reported model is in the biophysical mechanism detailing ionic and neutral solute transport for a large number of solutes, which were not taken into account in the earlier reported biophysical models of MS. Therefore, the reported model allows understanding response of mitochondria to the changes of initial concentration(s) of any of the solute(s) included in the model. Note that the values of all of the model parameters and kinetic constants have been estimated and the resulting complete model may be used for quantitative analysis of mitochondrial swelling dynamics in conditions of real in vitro experiments.

Stretching tension effects in permeability transition pores of inner mitochondrial membrane.

Presently a mechanism of permeability transition pore (PTP) opening was proposed and discussed. This mechanism is based on mechanical stretching of inner mitochondrial membrane (IMM) caused by mitochondrial swelling (MS). The latter is induced by osmotic pressure generated by solute imbalance between the matrix and the surrounding cyto(sarco)plasm. Modelled by the Monte-Carlo method, an IMM fragment of 350 simulated biological molecules exhibited formation of micro-domains containing two protein and seven phospholipid molecules. The energies (-0.191 eV per molecule) in these micro-domains were significantly larger than those (-0.375 eV per molecule) of other parts of the IMM fragment. Stretching forces applied to such domains expanded them much more than other parts of the IMM fragment. We identify these micro-domains as the PTPs. Both linear and nonlinear functions were used for the strain-stress relation of the IMM fragment, with nonlinear effects more important at large IMM stretching strains. Thus, two main factors are incorporated into the PTP opening mechanism: (1) presence of micro-domains in the IMM structure and (2) IMM stretching stress caused by MS. Taking into account both of these factors, the equation for the probability of PTP opening was deduced, with matrix Ca2+ and H+ ionic concentrations as its parameters. Note that the equation deduced was similar to an earlier reported empirical equation describing PTP opening dynamics. This correspondence provides support to the presently proposed mechanism. Thus, a new look at the PTP opening mechanism is provided, of interest to various research areas related to mitochondrial biophysics.

Reaction coupling in ADH1A alcohol dehydrogenase enzyme by exciplex formation with adenosine diphosphate moderated by low-energy electronic excited states.

Two commonly accepted theories about enzymes were revisited. The first states that adenosine triphosphate (ATP)-stored energy is only released when the substrate is in place, because the substrate changes the enzyme structure when it is bound to the enzyme. In fact, as demonstrated and discussed presently, no structural changes are required, and ATP-stored energy is released when it can be used. The second states that ATP-released energy moves along the enzyme molecule in the form of molecular vibrations (Davydov's vibrational solitons). In fact, as reported presently, energy released upon ATP hydrolysis moves in the form of excited-state electrons (excitons), with no molecular vibrations involved. The relevant experimental evidence was obtained for the human ADH1A alcohol dehydrogenase enzyme. Spontaneous ATP hydrolysis in the absence of substrate was apparently prevented by electronically excited enzyme + adenosine diphosphate (ADP) + inorganic phosphate (P) complex (exciplex) formed upon ATP hydrolysis. This exciplex kept ADP + P bound and in place for the inverse reaction, until the excess energy was dissipated in the enzyme-catalyzed reaction or by energy transfer to a suitable acceptor. Additionally, and contrary to textbooks, ADH1A has required ATP, working orders of magnitude faster in its presence.

Energy transfer along Müller cell intermediate filaments isolated from porcine retina: I. Excitons produced by ADH1A dimers upon simultaneous hydrolysis of two ATP molecules.

IR exciton propagation was explored in Müller cell (MC) intermediate filaments (IFs) filling a capillary matrix. These IFs have been isolated from porcine retina using different methods, while their properties were almost identical. Therefore, IFs isolated from the whole retinas were used presently. IR excitons were generated by IR radiation at 2 µm wavelength, or by enzymatic ATP hydrolysis, with the energy transferred to IFs. Excitons produced by ATP hydrolysis required simultaneous energy contribution of two ATP molecules, indicating simultaneous hydrolysis of two ATP molecules in the naturally dimeric human alcohol dehydrogenase enzyme (ADH1A). ATP hydrolysis was thus catalyzed by ADH1A…NAD+ enzymatic complexes absorbed at the IF extremities protruding out of the capillary matrix. The IR emission spectra of excitons were dependent on the exciton generation method. We believe this resulted from the exciton energy distribution varying in function of the generation method used. The latter seems reasonable, given the very long excited-state lifetimes, implying low nonradiative relaxation rates. The energy liberated by ATP hydrolysis has been measured directly in these experiments, for the first time. The results demonstrate that contrary to the predictions of equilibrium thermodynamics, the liberated energy is independent on the ATP/ADP concentration ratio, indicating that non-equilibrium reactions take place. Time-resolved experiments with excitons produced by pulsed IR radiation evaluated characteristic exciton propagation and emission times. For the first time, biexcitonic processes were observed in biological objects, whereby simultaneous hydrolysis of two ATP molecules bound to the same dimeric ADH1A molecule generated excitons carrying twice the energy liberated by hydrolysis of a single ATP molecule. The results reported indicate that ATP-liberated energy may be transmitted along natural polypeptide nanofibers in vivo, within and between live cells. These ideas could promote new understanding of the biophysics of life.

Supercritical Fluid Gaseous and Liquid States: A Review of Experimental Results.

We review the experimental evidence, from both historic and modern literature of thermodynamic properties, for the non-existence of a critical-point singularity on Gibbs density surface, for the existence of a critical density hiatus line between 2-phase coexistence, for a supercritical mesophase with the colloidal characteristics of a one-component 2-state phase, and for the percolation loci that bound the existence of gaseous and liquid states. An absence of any critical-point singularity is supported by an overwhelming body of experimental evidence dating back to the original pressure-volume-temperature (p-V-T) equation-of-state measurements of CO2 by Andrews in 1863, and extending to the present NIST-2019 Thermo-physical Properties data bank of more than 200 fluids. Historic heat capacity measurements in the 1960s that gave rise to the concept of "universality" are revisited. The only experimental evidence cited by the original protagonists of the van der Waals hypothesis, and universality theorists, is a misinterpretation of the isochoric heat capacity Cv. We conclude that the body of extensive scientific experimental evidence has never supported the Andrews-van der Waals theory of continuity of liquid and gas, or the existence of a singular critical point with universal scaling properties. All available thermodynamic experimental data, including modern computer experiments, are compatible with a critical divide at Tc, defined by the intersection of two percolation loci at gaseous and liquid phase bounds, and the existence of a colloid-like supercritical mesophase comprising both gaseous and liquid states.

Electron microscopy study of the central retinal fovea in Pied flycatcher: evidence of a mechanism of light energy transmission through the retina.

We present unique ultrastructural data on avian retinal cells. Presently and earlier (Zueva et al., 2016) we explored distribution of intermediate filaments (IFs) in retinal cells of the Pied flycatcher (Ficedula hypoleuca, Passeriformes, Aves) in the central foveolar zone. This retinal zone only contains single and double cone photoreceptors. Previously we found that continuous IFs span Müller cells (MC) lengthwise from the retinal inner limiting membrane (ILM) layer up to the outer limiting membrane (OLM) layer. Here we describe long cylindrical bundles of IFs (IFBs) inside the cone inner segments (CIS) adjoining the cone plasma membrane, with these IFBs following along the cone lengthwise, and surrounding the cone at equal spacing one from the other. Double cones form a combined unit, wherein they are separated by their respective plasma membranes. Double cones thus have a common external ring of IFBs, surrounding both cone components. In the layer of cilia, the IFBs that continue into the cone outer segment (COS) follow on to the cone apical tip along the direction of incident light, with single IFs separating from the IFB, touching, and sometimes passing in-between the light-sensitive lamellae of the COS. These new data support our previous hypothesis on the quantum mechanism of light energy propagation through the vertebrate retina (Zueva et al., 2016, 2019).

Absorption spectra of Müller cell intermediate filaments: Experimental results and theoretical models.

Experimental spectra of Müller cell (MC) intermediate filaments (IFs) isolated from porcine retina are reported in this work. The absorption spectra recorded at different MC IF concentrations were used to estimate their absorption cross-sections at different wavelengths. The average absorption cross-section of a single MC IF was ca. (0.97 … 2.01) × 10-10 cm2 in the 650-445 nm spectral range. To interpret these experimental absorption spectra, we made ab initio calculations of the optical spectra of α-helix polypeptides, and also used a simplified theoretical approach that modeled an IF by a conductive wire. The energy spectra of the refractive index, extinction coefficient (absorption cross-section), energy loss and reflectivity functions for different photon polarizations, with strong anisotropy with respect to the system axis, were calculated ab initio for polyglycine α-helix molecule containing 1000 glycine residues. Strong anisotropy of these parameters was explained by photons interacting with different electronic transitions. Note that similarly strong anisotropy was also obtained for the optical absorption cross-sections in the simplified model. Both modeling approaches were used for calculating the absorption cross-sections of interest. As a result, the absorption cross-section for photons propagating axially along MC IFs was much larger than their geometrical cross-section. The latter result was explained taking into account the density of electronic states, with numerous electrons contributing to the transition intensity at a given energy. We found that the simple conductive wire model describes the MC IF absorption spectrum better than the ab initio spectra. The latter conclusion was explained by the limitations of ab initio analysis, which only took into account one α-helix with 1000 aminoacids, whereas each porcine Müller cell IF is assembled of thousands of protein molecules, reaching the total length of ca. 100 µm. The presently reported results contribute to the understanding of the quantum mechanism of high-contrast vision of vertebrate eyes.

On the Effects of Mechanical Stress of Biological Membranes in Modeling of Swelling Dynamics of Biological Systems.

We highlight mechanical stretching and bending of membranes and the importance of membrane deformations in the analysis of swelling dynamics of biological systems, including cells and subcellular organelles. Membrane deformation upon swelling generates tensile stress and internal pressure, contributing to volume changes in biological systems. Therefore, in addition to physical (internal/external) and chemical factors, mechanical properties of the membranes should be considered in modeling analysis of cellular swelling. Here we describe an approach that considers mechanical properties of the membranes in the analysis of swelling dynamics of biological systems. This approach includes membrane bending and stretching deformations into the model, producing a more realistic description of swelling. We also discuss the effects of membrane stretching on swelling dynamics. We report that additional pressure generated by membrane bending is negligible, compared to pressures generated by membrane stretching, when both membrane surface area and volume are variable parameters. Note that bending deformations are reversible, while stretching deformation may be irreversible, leading to membrane disruption when they exceed a certain threshold level. Therefore, bending deformations need only be considered in reversible physiological swelling, whereas stretching deformations should also be considered in pathological irreversible swelling. Thus, the currently proposed approach may be used to develop a detailed biophysical model describing the transition from physiological to pathological swelling mode.

Application of multidimensional and conventional fluorescence techniques for classification of beverages originating from various berry fruit.

The objectives of this study were to characterize fluorescence of beverages from berry fruit, including chokeberry, blackcurrant, raspberry and strawberry, and to develop classification models based on different types of fluorescence spectra to identify beverages depending on the fruit species. Total fluorescence spectra (excitation-emission matrices, EEMs) and total synchronous fluorescence spectra (TSFS) were recorded for a series of commercial berry fruit beverages. An analysis of EEMs using parallel factor analysis (PARAFAC) revealed four components characterized by the excitation/emission maxima at 275/326, 319/410, 414/600, and 360/460 nm, respectively. Based on the spectral profiles, these components were assigned to various groups of phenolic compounds. Partial least squares discriminant analysis was used to develop the classification models. The analysis was performed on PARAFAC scores, unfolded EEMs (uEEMs), unfolded TSFS (uTSFS), and additionally on conventional emission spectra (EMS) measured at particular excitation wavelengths and single synchronous fluorescence spectra (SFS). The classification models with the same average classification error of 4.86% were obtained for the analysis of both the entire uEEMs and uTSFS. Among models based on the individual spectra, the lowest error of 4.42% was obtained for SFS measured at Δλ = 40 nm, and an error of 7.64% was obtained for EMS measured at the excitation wavelength of 360 nm. The classification model based on the PARAFAC scores had the highest error of 15.27%. The present results show good potential of fluorescence as rapid and reagent-free tool for authenticity evaluation of berry beverages.