HNCA-TOCSY-CANH experiments with alternate <Superscript>13</Superscript>C-<Superscript>12</Superscript>C labeling: a set of 3D experiment with unique supra-sequential information for mainchain resonance assignment

Described here is a set of three-dimensional (3D) NMR experiments that rely on CACA-TOCSY magnetization transfer via the weak ³ \textJ_{\textCa\textCa} ^{ 3} {\text{J}}_{{{\text{C}}\alpha {\text{C}}\alpha }} coupling. These pulse sequences, which resemble recently described ¹³C detected CACA-TOCSY (Takeuchi et al. 2010) experiments, are recorded in ¹H₂O, and use ¹H excitation and detection. These experiments require alternate ¹³C-¹²C labeling together with perdeuteration, which allows utilizing the small ³ \textJ_{\textCa\textCa} ^{ 3} {\text{J}}_{{{\text{C}}\alpha {\text{C}}\alpha }} scalar coupling that is otherwise masked by the stronger ¹J_CC couplings in uniformly ¹³C labeled samples. These new experiments provide a unique assignment ladder-mark that yields bidirectional supra-sequential information and can readily straddle proline residues. Unlike the conventional HNCA experiment, which contains only sequential information to the ^{1 3} \textC^a ^{ 1 3} {\text{C}}^{\alpha } of the preceding residue, the 3D hnCA-TOCSY-caNH experiment can yield sequential correlations to alpha carbons in positions i−1, i + 1 and i−2. Furthermore, the 3D hNca-TOCSY-caNH and Hnca-TOCSY-caNH experiments, which share the same magnetization pathway but use a different chemical shift encoding, directly couple the ¹⁵N-¹H spin pair of residue i to adjacent amide protons and nitrogens at positions i−2, i−1, i + 1 and i + 2, respectively. These new experimental features make protein backbone assignments more robust by reducing the degeneracy problem associated with the conventional 3D NMR experiments.     

Influence of {\text{NH - }}{{\text{S}}^\gamma } bonding interactions on the structure and dynamics of metallothioneins

Mammalian metallothioneins ( \textM₇^\textIIMTs {\text{M}}_7^{\text{IIMTs}} ) show a clustered arrangement of the metal ions and a nonregular protein structure. The solution structures of Cd₃-thiolate cluster containing β-domain of mouse β-MT-1 and rat β-MT-2 show high structural similarities, but widely differing structure dynamics. Molecular dynamics simulations revealed a substantially increased number of \textNH - \textS^g {\text{NH - }}{{\text{S}}^\gamma } hydrogen bonds in β-MT-2, features likely responsible for the increased stability of the Cd₃-thiolate cluster and the enfolding protein domain. Alterations in the \textNH - \textS^g {\text{NH - }}{{\text{S}}^\gamma } hydrogen-bonding network may provide a rationale for the differences in dynamic properties encountered in the β-domains of MT-1, -2, and -3 isoforms, believed to be essential for their different biological function.     

Genetic parameters and predicted selection responses for timber production traits in a Castanea sativa progeny trial: developing a breeding program

Total height, diameter, index volume, stem straightness, apical dominance, and survival were assessed at 8 years from seed in an open-pollinated progeny test of 36 families of European chestnut (Castanea sativa Miller) established at two sites in the Atlantic area of Galicia, Spain. Iterative spatial analysis was applied to eliminate the effect of the spatial dependence in the original data and to estimate accurately genetic parameters for evaluating the potential for selection of the measured trees. Spatial analysis was very beneficial for growth traits and survival, but less so if at all for form traits. Estimated individual heritabilities ranged from moderate to high for growth traits ([^(h)]_i² = 0.29 - 0.42 \widehat{h}_i^2 = 0.29 - 0.42 ) and stem straightness ([^(h)]_i² = 0.24 - 0.42 \widehat{h}_i^2 = 0.{24} - 0.{42} ). High coefficients of additive genetic variance were obtained for volume ( [^(\textC)]\textV_\textA = 36.5 - 41.5% \widehat{\text{C}}{{\text{V}}_{\text{A}}} = {36}.{5} - {41}.{5}\% ) and straightness ( [^(\textC)]\textV_\textA = 44.26 - 53.84% \widehat{\text{C}}{{\text{V}}_{\text{A}}} = {44}.{26} - {53}.{84}\% ). Phenotypic and estimated genetic correlations between growth traits were very high, and correlations between sites indicated that there was no important family × site interaction. No adverse correlations between traits were evident. The results indicate the ample potential for selection in the current progeny trial, where responses to within-family and combined selection for growth traits may be high. Accordingly, three selection scenarios were addressed with the aim to initiate the selection of individuals for implementing the Forest Breeding Plan of Galicia for European chestnut.     

Parameterization of chlorophyll a-specific absorption coefficients and effects of their variations in a highly eutrophic lake: a case study at Lake Kasumigaura, Japan

The chlorophyll a-specific absorption coefficient ( a_\textph^* ( l) a_{\text{ph}}^{*} \left( \lambda \right) ) in a highly eutrophic lake can show characteristics distinct from that in the ocean due to the differences in the structure and composition of phytoplankton. In this study, investigated the variation of a_\textph^* ( l) a_{\text{ph}}^{*} \left( \lambda \right) in Lake Kasumigaura, a highly eutrophic lake in Japan, in association with the package effect and the effect of accessory pigments, and carried out the parameterization of a_\textph^* ( l) a_{\text{ph}}^{*} \left( \lambda \right) . Although a_\textph^* ( l) a_{\text{ph}}^{*} \left( \lambda \right) did not vary spatially, it did show significant temporal variation, with a particularly high value after spring-bloom. This high a_\textph^* ( l) a_{\text{ph}}^{*} \left( \lambda \right) in spring was attributed to a lower package effect and a higher proportion of carotenoid than the other samples. Although the value of a_\textph^* ( l) a_{\text{ph}}^{*} \left( \lambda \right) was correlated with the concentration of chlorophyll-a (Chl-a), the correlation coefficient was lower than those reported in the ocean. Some lake-water samples showed variations of the package effect and the effect of accessory pigments that were independent of the concentration of Chl-a, and these independent variations resulted in the weak correlation between a_\textph^* ( l) a_{\text{ph}}^{*} \left( \lambda \right) and the concentration of Chl-a. Together, these results suggest that the factors controlling a_\textph^* ( l) a_{\text{ph}}^{*} \left( \lambda \right) in highly eutrophic lakes are distinct from that in ocean samples.     

3D-QSAR studies of triazolopyrimidine derivatives of <Emphasis Type="Italic">Plasmodium falciparum</Emphasis> dihydroorotate dehydrogenase inhibitors using a combination of molecular dynamics,docking, and genetic algorithm-based methods

A series of 35 triazolopyrimidine analogues reported as Plasmodium falciparum dihydroorotate dehydrogenase (PfDHODH) inhibitors were optimized using quantum mechanics methods, and their binding conformations were studied by docking and 3D quantitative structure–activity relationship studies. Genetic algorithm-based criteria was adopted for selection of training and test sets while maintaining structural diversity of training and test sets, which is also very crucial for model development and validation. Both the comparative molecular field analyses ( q_\textLOO² = 0.841 q_{\text{LOO}}^2 = 0.{841} , r_\textncv² = 0.99 r_{\text{ncv}}^2 = 0.{99} ) and comparative molecular similarity indices analyses ( q_\textLOO² = 0.757 q_{\text{LOO}}^2 = 0.{757} , r_\textncv² = 0.943 r_{\text{ncv}}^2 = 0.{943} ) show excellent correlation and high predictive power. Furthermore, molecular dynamics simulations were performed to explore the binding mode of the two of the most active compounds of the series, 10 and 14. Harmonization in the two simulation results validate the analysis and therefore applicability of docking parameters based on crystallographic conformation of compound 14 bound to receptor molecule. This work provides useful information about the inhibition mechanism of this class of molecules and will assist in the design of more potent inhibitors of PfDHODH.     

Strategies for efficient repetitive production of succinate using metabolically engineered Escherichia coli

Escherichia coli AFP111, a pflB, ldhA, ptsG triple mutant of E. coli W1485, can be recovered for additional succinate production in fresh medium after two-stage fermentation (an aerobic growth stage followed by an anaerobic production stage). However, the specific productivity is lower than that of two-stage fermentation. In this study, three strategies were compared for reusing the cells. It was found when cells were aerobically cultivated at the end of two-stage fermentation without supplementing any carbon source, metabolites (mainly succinate and acetate) could be consumed. As a result, enzyme activities involved in the reductive arm of tricarboxylic acid cycle and the glyoxylate shunt were enhanced, yielding a succinate specific productivity above 1 2 5  \textmg  \textg_DCW ^{- 1}  \texth ^{- 1} 1 2 5\;{\text{mg}}\;{\text{g}}_{\rm DCW}^{ - 1} \,{\text{h}}^{ - 1} and a mass yield above 0.90 g g⁻¹ in the subsequent anaerobic fermentation. In addition, the intracellular NADH of cells subjected to aerobic cultivation with metabolites increased by more than 3.6 times and the ratio of NADH to NAD⁺ increased from 0.4 to 1.3, which were both favorable for driving the TCA branch to succinate.     

A standard quantitative method to measure acid tolerance of probiotic cells

The aim of this work was to develop a standard quantitative method to measure the acid tolerance of probiotic cells when exposed to a simulated gastric fluid. Three model strains of different cell concentrations were exposed to a standard simulated gastric fluid of fixed volume. The fluid pH ranged from pH 1.5 to 2.5. In general, the death kinetics followed an exponential trend. The overall death constant, k _d, for all strains was found to be in a power relationship with the pH value and the initial cell concentration, and it can be expressed as

Genetic effects of forest fragmentation in high-density Araucaria angustifolia populations in Southern Brazil

Araucaria angustifolia is an endangered tropical/subtropical coniferous of great interest for conservation due its economical, ecological, and social value. Only 3% of original Araucaria forests remain, which are generally confined to small forest fragments. Forest fragmentation can have serious consequences on genetic process in tree population, affecting long-term fitness and adaptability. To investigate the effects of forest fragmentation on genetic diversity and the structure of A. angustifolia populations, the genetic diversity of eight microsatellite loci was compared in four small fragmented populations (<22 ha), four tree groups (five to 11 trees) occurring in pastures and in three plots in a large continuous population. The clearest effect of fragmentation was the loss of rare alleles (p ≤ 0.05) in fragmented populations (19.4% to 47.2%) and intermediate frequency (0.05 < p ≤ 0.25) and rare alleles (p ≤ 0.05) in tree groups (19% to 86.1%) in comparison to continuous populations. Fragmented populations have significant higher fixation index ( [^(F)]_\textIS = 0.121 \widehat{F}_{\text{IS}} = 0.121 , P < 0.05) than continuous populations ( [^(F)]_\textIS = 0.083 \widehat{F}_{\text{IS}} = 0.083 , P < 0.05). High genetic differentiation was detected among tree groups ( [^(G)]_\textST^￠ = 0.258 \widehat{G}_{{{\text{ST}}}}^{\prime } = 0.258 , P < 0.01) and low among fragments ( [^(G)]_\textST^￠ = 0.031 \widehat{G}_{{{\text{ST}}}}^{\prime } = 0.031 , P < 0.05) and continuous populations ( [^(G)]_\textST^￠ = 0.026 \widehat{G}_{{{\text{ST}}}}^{\prime } = 0.026 , P < 0.05), showing a significant bottleneck effect in tree groups. Evidence that forest fragments have experienced a recent bottleneck was confirmed in at least two studied fragments. The implications of the results for conservation of the fragmented A. angustifolia populations are discussed.     

Physiological characterization of ‘stay green’ wheat cultivars during the grain filling stage under field growing conditions

Rye (Secale cereale L.) chromosome arm 1RS could delay leaf senescence, and change in H₂O₂ content is a useful index for weighing the ability to delay the senescence. Two wheat cultivars, Chuannong12 (CN12) and Chuannong 18 (CN18), harboring the wheat–rye 1BL/1RS translocated chromosome were investigated for H₂O₂ change and physiological index after flowering under field conditions, and MY11, the agronomical parent of both CN12 and CN18, was used as the control. A combined change in the peak value of CdSe/ZnS quantum dot (QD) fluorescence and morphological observation indicated that the H₂O₂ contents in CN12 and CN18 were generally lower than that in MY11. They both had higher values for net photosynthetic rate (P _n), stomatal conductance (G _s), F_\textv /F_\textm^￠ F_{\text{v}} /F_{\text{m}}^{\prime } F_\textv^￠ /F_\textm^￠ F_{\text{v}}^{\prime } /F_{\text{m}}^{\prime } , and photochemical quenching of PSII (qP) than MY11 only in the late measurement stage. Some small differences were also observed, such as CN12 and CN18 wheat cultivars having higher and longer photosynthetic competence than MY11 during the grain filling stage, which perhaps resulted from a mechanism for removing oxidative species, especially H₂O₂.     

Effects of nitrate on intracellular nitrite and growth of Microcystis aeruginosa

Although nitrate is a macronutrient and can serve as good nitrogen source for many species of phytoplankton, high nitrate concentrations do not benefit the growth of phytoplankton. We hypothesise that algae cultured under high nitrate concentrations can accumulate intracellular nitrite, which is produced by nitrate reductase (NR) and can inhibit the growth of algae. To assess the validity of this hypothesis, Microcystis aeruginosa was grown under different nitrate concentrations from 3.57 to 21.43 mM in low CO₂ and high CO₂ conditions for 15 days. We observed that, with increasing nitrate concentrations, the intracellular nitrite concentrations of the alga increased and the growth rates and photosynthesis declined. When grown under high CO₂ conditions, M. aeruginosa showed lower intracellular nitrite concentrations and higher growth rates and \textP_\textm^\textchla {\text{P}}_{\text{m}}^{{\text{chl}}a} , \textR_\textd^\textchla {\text{R}}_{\text{d}}^{{\text{chl}}a} , α^chla than under low CO₂ conditions. These results suggest that the accumulation of intracellular nitrite could be the cause of inhibition of algal growth under high nitrate concentrations.     

A step-forward in the characterization and potential applications of solid and liquid oxygen transfer vectors

Silicone oil 20 and 200 cSt, a perfluorocarbon (FC40TM), heptamethylnonane, Kraton, Elvax, and Desmopan were evaluated for their ability to enhance oxygen transfer in stirred tank and airlift reactors (STR and ALR, respectively). None of the vectors tested was either toxic or biodegradable and they exhibited a moderate affinity for oxygen (gas/vector partitioning coefficients K_{\textg/\textv} = C_\textg ·C_\textv^{- 1} K_{{{\text{g}}/{\text{v}}}} = C_{\text{g}} \cdot C_{\text{v}}^{- 1} ranging from 3 to 5.1). FC40 was highly volatile, while KratonTM and ElvaxTM exhibited a low thermal stability, which constitutes a serious handicap for their implementation in fermentations. Silicone oil 20 cSt and Desmopan supported the highest oxygen transfer rates under abiotic conditions in both STR and ALR designs, with enhancement factors of up to 90% and 250%, respectively, compared to control tests (deprived of vector). The fact that these vectors showed the highest K _g/v proved that, besides the classical selection criteria, the in situ hydrodynamic behavior (which affects K _L a) must be considered for vector selection. The use of silicone oil 20 cSt and Desmopan in glucose-supplemented Saccharomyces cerevisiae fermentations resulted in a two- and threefold increase in biomass productions, respectively. The better performance of Desmopan in terms of biomass growth enhancement, together with the absence of the operational problems inherent to the use of liquid vectors (such as intensive foaming, high cost, and difficult solvent recovery), make solid vectors a promising and cost-effective alternative in the future developments of two-phase partitioning bioreactors.     

Kinetics of styrene biodegradation by <Emphasis Type="Italic">Pseudomonas</Emphasis> sp. E-93486

The research into kinetics of styrene biodegradation by bacterial strain Pseudomonas sp. E-93486 coming from VTT Culture Collection (Finland) was presented in this work. Microbial growth tests in the presence of styrene as the sole carbon and energy source were performed both in batch and continuous cultures. Batch experiments were conducted for initial concentration of styrene in the liquid phase changed in the range of 5–90 g m⁻³. The Haldane model was found to be the best to fit the kinetic data, and the estimated constants of the equation were: μ _m = 0.1188 h⁻¹, K _S = 5.984 mg l⁻¹, and K _i = 156.6 mg l⁻¹. The yield coefficient mean value Y_\textxs^\textapp Y_{\text{xs}}^{\text{app}} for the batch culture was 0.72 g_{dry cells weight} (g_substrate)⁻¹. The experiments conducted in a chemostat at various dilution rates (D = 0.035–0.1 h⁻¹) made it possible to determine the value of the coefficient for maintenance metabolism m _d = 0.0165 h⁻¹ and the maximum yield coefficient value Y_\textxs^\textM = 0.913 Y_{\text{xs}}^{\text{M}} = 0.913 . Chemostat experiments confirmed the high value of yield coefficient Y_\textxs^\textapp Y_{\text{xs}}^{\text{app}} observed in the batch culture. The conducted experiments showed high activity of the examined strain in the styrene biodegradation process and a relatively low sensitivity to inhibition of its growth at higher concentrations of styrene in the solution. Such exceptional features of Pseudomonas sp. E-93486 make this bacterial strain the perfect candidate for technical applications.     

Toxicity assessment of common organic solvents using a biosensor based on algal photosynthetic activity measurement

The acute toxicities of common organic solvents (e.g., methanol, ethanol, isopropanol, acetone, acetonitrile, and dimethylformamide) were evaluated using a biosensor based on microalgal photosynthesis measurement. The biosensor was air-tight, with no headspace, preventing volatile organic toxicants from escaping into the environment as well as partitioning from the aqueous phase into the headspace until equilibrium was reached. Both the incubating and exposure times were set at 10 min. It was observed that only 2 h was needed to obtain complete dose-related inhibition of photosynthetic activity. The results showed that all the tested organic solvents inhibited algal photosynthesis with EC₅₀ ranging between 589 and 2,570 mM. The inhibition of these solvents was in the order: isopropanol > acetone > acetonitrile > ethanol > dimethylformamide > methanol. The quantitative structure-activity relationship (QSAR) between toxicity data and partition coefficient of the examined compounds could be modeled as follows: ${\text{log}}_{{10}} {\text{EC}}_{{50}} \;{\left( {\mu {\text{M}}} \right)} = - 0.6428\;{\text{log}}\;P + 5.76\;{\left( {{\text{R}}^{2} \approx 0.88} \right)}The acute toxicities of common organic solvents (e.g., methanol, ethanol, isopropanol, acetone, acetonitrile, and dimethylformamide) were evaluated using a biosensor based on microalgal photosynthesis measurement. The biosensor was air-tight, with no headspace, preventing volatile organic toxicants from escaping into the environment as well as partitioning from the aqueous phase into the headspace until equilibrium was reached. Both the incubating and exposure times were set at 10 min. It was observed that only 2 h was needed to obtain complete dose-related inhibition of photosynthetic activity. The results showed that all the tested organic solvents inhibited algal photosynthesis with EC₅₀ ranging between 589 and 2,570 mM. The inhibition of these solvents was in the order: isopropanol > acetone > acetonitrile > ethanol > dimethylformamide > methanol. The quantitative structure-activity relationship (QSAR) between toxicity data and partition coefficient of the examined compounds could be modeled as follows: \textlog₁₀ \textEC₅₀   ( m\textM ) = - 0.6428  \textlog  P + 5.76  ( \textR² ? 0.88 ){\text{log}}_{{10}} {\text{EC}}_{{50}} \;{\left( {\mu {\text{M}}} \right)} = - 0.6428\;{\text{log}}\;P + 5.76\;{\left( {{\text{R}}^{2} \approx 0.88} \right)}. This indicates that the photosynthetic activity of the microalga Pseudokirchneriella subcapitata is highly dependent on the hydrophobicity of these commonly used organic solvents.     

Thermal radiation absorbed by dairy cows in pasture

The goal of the present paper was to assess a method for estimating the thermal radiation absorbed by dairy cows (0.875 Holstein–0.125 Guzerath) on pasture. A field test was conducted with 472 crossbred dairy cows in three locations of a tropical region. The following environmental data were collected: air temperature, partial vapour pressure, wind speed, black globe temperature, ground surface temperature and solar radiation. Average total radiation absorbed by animals was calculated as R_abs = 640.0 ±3.1 W.m ^{- 2} {R_{abs}} = 640.0 \pm 3.1\, W.{m^{ - 2}} . Absorbed short-wave radiation (solar direct, diffuse and reflected) averaged 297.9 ± 2.7 W m⁻²; long wave (from the sky and from terrestrial surfaces) averaged 342.1 ± 1.5 W m⁻². It was suggested that a new environmental measurement, the effective radiant heat load (ERHL), could be used to assess the effective mean radiant temperature ( T_\textmr^* ) \left( {T_{\text{mr}}^* } \right) . Average T_\textmr^* T_{\text{mr}}^* was 101.4 ± 1.2°C, in contrast to the usual mean radiant temperature, T_mr = 65.1 ±0.5^° C {T_{mr}} = 65.1 \pm 0.5^\circ C . Estimates of T_\textmr^* T_{\text{mr}}^* were considered as more reliable than those of T _mr in evaluating the thermal environment in the open field, because T _mr is almost totally associated only with long wave radiation.     

Analysis of the activation mechanism of <Emphasis Type="Italic">Pseudomonas stutzeri</Emphasis> cytochrome <Emphasis Type="Italic">c</Emphasis> peroxidase through an electron transfer chain

The activation mechanism of Pseudomonas stutzeri cytochrome c peroxidase (CCP) was probed through the mediated electrochemical catalysis by its physiological electron donor, P. stutzeri cytochrome c-551. A comparative study was carried out, by performing assays with the enzyme in the resting oxidized state as well as in the mixed-valence activated form, using cyclic voltammetry and a pyrolytic graphite membrane electrode. In the presence of both the enzyme and hydrogen peroxide, the peak-like signal of cytochrome c-551 is converted into a sigmoidal wave form characteristic of an \textE_\textr \textC_\texti^￠ {\text{E}}_{\text{r}} {\text{C}}_{\text{i}}^{\prime } catalytic mechanism. An intermolecular electron transfer rate constant of (4 ± 1) × 10⁵ M⁻¹ s⁻¹ was estimated for both forms of the enzyme, as well as a similar Michaelis–Menten constant. These results show that neither the intermolecular electron transfer nor the catalytic activity is kinetically controlled by the activation mechanism of CCP in the case of the P. stutzeri enzyme. Direct enzyme catalysis using protein film voltammetry was unsuccessful for the analysis of the activation mechanism, since P. stutzeri CCP undergoes an undesirable interaction with the pyrolytic graphite surface. This interaction, previously reported for the Paracoccus pantotrophus CCP, induces the formation of a non-native conformation state of the electron-transferring haem, which has a redox potential 200 mV lower than that of the native state and maintains peroxidatic activity.     

A molecular dynamics study on opioid activities of biphalin molecule

Molecular dynamics simulations of the biphalin molecule, (Tyr-D-Ala-Gly-Phe-NH)₂, and the active tetrapeptide hydrazide, Tyr-D-Ala-Gly-Phe-NH-NH₂ were performed to investigate the cause of the increased μ and δ receptor binding affinities of the former over the latter. The simulation results demonstrate that the acylation of the two equal tetrapeptide fragments of biphalin produces the constrained hydrazide bridges C₄^a - C₄￠- N₉ - N₁₀ {\hbox{C}}_4^{\alpha } - {{\hbox{C}}_4}\prime - {{\hbox{N}}_9} - {{\hbox{N}}_{{10}}} and N₉ - N₁₀ - C₅￠- C₅^a {{\hbox{N}}_9} - {{\hbox{N}}_{{10}}} - {{\hbox{C}}_5}\prime - {\hbox{C}}_5^{\alpha } , which in turn increase the opportunity of conformations for binding to μ or δ receptors. Meanwhile, the connection of the two active tetrapeptide fragments of biphalin also results in the constrained side chain torsion angle χ² at one of the two residues Phe. This constrained side chain torsion angle not only significantly increases the δ receptor binding affinity but also makes most of the δ receptor binding conformations of biphalin bind to the δ receptor through the fragment containing the mentioned residue Phe.     

The Equilibria of Lipid–K<Superscript>+</Superscript> Ions in Monolayer at the Air/Water Interface

The effect of K⁺ ion interaction with monolayers of phosphatidylcholine (lecithin, PC) or cholesterol (Ch) was investigated at the air/water interface. We present surface tension measurements of lipid monolayers obtained using a Langmuir method as a function of K⁺ ion concentration. Measurements were carried out at 22°C using a Teflon trough and a Nima 9000 tensiometer. Interactions between lecithin and K⁺ ions or Ch and K⁺ ions result in significant deviations from the additivity rule. An equilibrium theory to describe the behavior of monolayer components at the air/water interface was developed in order to obtain the stability constants and area occupied by one molecule of lipid–K⁺ ion complex (LK⁺). The stability constants for lecithin–K⁺ ion (PCK⁺) complex, \( K_{{{\text{PCK}}^{ + } }} = { 3}. 2 6\times 10^{ 2} {\text{dm}}^{ 3} \,{\text{mol}}^{ - 1} \), and for cholesterol–K⁺ ion (ChK⁺) complex, \( K_{{{\text{ChK}}^{ + } }} = { 1}.00 \times 10^{ 3} {\text{dm}}^{ 3} \,{\text{mol}}^{ - 1} \), were calculated by inserting the experimental data. The value of area occupied by one PCK⁺ complex is 60 Å² molecule^?1, while the area occupied by one ChK⁺ complex is 40.9 Å² molecule^?1. The complex formation energy (Gibbs free energy) values for the PCK⁺ and ChK⁺ complexes are ?14.18 ± 0.71 and ?16.92 ± 0.85 kJ mol^?1, respectively.