Prediction by computer modelling of the precipitation of stone-forming solids from urine

The development and verification of an equilibrium speciation computer model of urine is described. The model is based on critically selected equilibrium constants and solubility products corrected where needed to T=37°C and I=0.2 mol dm⁻³. The model represents a slight improvement over previous equilibrium models in that a simpler and more reliable measure is used to represent the degree of supersaturation with respect to a stone-forming salt. A further improvement arises from an implicit allowance being made for certain kinetic factors governing the precipitation of calcium-phosphate solids. The model was verified by comparison with an analogous experimental procedure applied to a standard reference artificial urine. Good predictions were obtained for the precipitation of calcium oxalate monohydrate (COM), dicalcium phosphate dihydrate (DCPD), calcium hydroxyapatite (HAP) and uric acid, but not for struvite. The model indicates that normal urine in the physiological pH range of 5.5 to 7.0 contains precipitates of COM, DCPD and HAP. Precipitates of uric acid and struvite are predicted to occur below and above a pH of 6, respectively. Further computations yield results which are consistent with the view that DCPD may act as a precursor in the precipitation of HAP. Finally, the results indicate that COM precipitation is far more markedly augmented by an increase in oxalate concentration than by an increase in calcium concentration.     

Photosynthesis,Respiration and Exopolymer Calcium-Binding in Biofilm Calcification (Westerhöfer and Deinschwanger Creek,Germany)

The impact of microbial activity on biofilm calcification in aquatic environments is still a matter of debate, especially in settings where ambient water has high CaCO₃ mineral supersaturation. In this study, biofilms of two CO₂-degassing karst-water creeks in Germany, which attain high calcite supersaturation during their course downstream, were investigated with regard to water chemistry of the biofilm microenvironment. The biofilms mainly consisted of filamentous cyanobacteria (Phormidium morphotype) and heterotrophic bacteria (including sulfate-reducing bacteria), which affect the microenvironment and produce acidic exopolymers. In situ and ex situ microelectrode measurements showed that a strong pH increase, coupled with Ca^{2 +} consumption, occurred in light conditions at the biofilm surface, while the opposite occurred in the dark. Calcite supersaturation at the biofilm surface, calculated from ex situ Ca^{2 +} and CO₃ ^2? microelectrode measurements, showed that photosynthesis resulted in high omega values during illumination, while respiration slightly lowered supersaturation values in the dark, compared to values in the water column. Dissociation calculation demonstrated that the potential amount of Ca^{2 +} binding by exopolymers would be insufficient to explain the Ca^{2 +} loss observed, although Ca^{2 +} complexation to exopolymers might be crucial for calcite nucleation. No spontaneous precipitation occurred on biofilm-free limestone substrates under the same condition, regardless of high supersaturation. These facts indicate that photosynthesis is a crucial mechanism to overcome the kinetic barrier for CaCO₃ precipitation, even in highly supersaturated settings.     

The biology of carbonate precipitation by cyanobacteria

Summary In the freshwater areas of the Everglades, Florida, U.S.A., carbonate is precipitated in dense cyanobacterial mats. Precipitation is linked with photosynthesis in the mats in a quantitative relationship. On ground of field observations and experiments a model for precipitation in the filamentous cyanobacteriaScytonema is proposed, which links precipitation to bicarbonate use in photosynthesis and subsequent release of OH⁻ ions. Besides supersaturation of the water with respect to carbonate and photosynthetic bicarbonate use, precipitation requires a suitable sheath structure and composition. The characteristics of the sheath seem to be responsible for a distinct crystal morphology in the two generaScytonema andSchizothrix, as well as for the restriction of calcification to the outer sheath inScytonema. In the immediate vicinity of the trichom precipitation seems to be inhibited. Comparison of this form of calcifying cyanobacteria with calcification in calcareous algae shows many similarities and rises the question of the biological significance of calcification or precipitation. The precipitated carbonate shows equilibrium precipitation in its δ oxygen values, while it is enriched in¹³C relative to the ambient water. This agrees with a model of precipitation in which the carbonate derives from the water immediately surrounding the filament. There the water is depleted in¹²C which is preferably taken up for photosynthesis. No respiratory carbon is involved in precipitation. From measurements of the amount of precipitation in the field and in experiments the annual sedimentation rate is estimated to be 0.024 to 0.24 mm. These values fall within the range of laminae thicknesses in fossil algal laminites.     

Sensitivity of the woodland herb Anemone hepatica to changing environmental conditions

Abstract. The decline of deciduous woodland populations of Anemone hepatica L. in southern Sweden is documented and possible causes are discussed. The study was based on (1) re‐investigations of 6.25 km² grid‐squares first studied in 1938–1970, (2) distribution of A. hepatica in woodland sites with well‐known soil chemical properties and (3) a detailed study over 12 consecutive years into the relationships between biological characteristics of the species (number of individuals, vegetative development, flowering frequency) and environmental variables (temperature, precipitation), soil chemistry and time. There was a close relationship between soil acidity (pH, solubility of Al³⁺) and both distribution and biological characteristics. The biological variables declined significantly over time but were not related to climatic variability. Increasing soil acidity and Al³⁺ solubility are concluded to be the main factors responsible for the decline of A. hepatica in S. Swedish deciduous woodlands.     

Quantitative analysis of drug-receptor interactions: I. Determination of kinetic and equilibrium properties

It has recently become possible to characterize a variety of different receptors by studying the binding of appropriate drugs labelled with ³H or ¹²⁵I. The goal of this review is to describe the basic mathematical analyses that should be used to characterize a particular receptor in terms of its interactions with ligands. Methods for direct determination of kinetic and equilibrium constants of simple bimolecular drug-receptor interactions are described as is the use of these measurements to verify the existence of a simple second order reaction. Some of the causes of deviations from second order behavior which imply more complex interactions are also discussed. $\text{In vitro}$ studies of radioligand binding provide a means of indirectly determining equilibrium dissociation constants of unlabelled drugs. The appropriate equations for these determinations are presented and the assumptions underlying these calculations are identified. Analysis of the temperature dependence of kinetic and equilibrium constants allows determination of the energetics of binding and methods are presented for calculation of the changes in Gibbs free energy, enthalpy and entropy that are associated with the binding of ligands to receptors. Studies of the interactions of agonists and antagonists with β-adrenergic receptors are presented as examples of the various types of calculations.     

A mathematical theory of enamel solubility and the onset of dental caries: II. Some solubility equilibrium considerations of hydroxyapatite

Equilibrium solubility considerations are presented based on the assumption that equating the kinetic expressionq, developed in part I, to zero can describe the equilibrium or steady state between hydroxyapatite and salt solutions. From this expression is derived Hodge's empirical equilibrium equation,C=KH. Further, a lograithmic transformation of this equation results in an expression that accounts for the equilibrium calcium, phosphorus andpH relation found by Levinskas and Neuman. Finally, it also shows the relation between log (C·P) andpH necessary for typical artificial carious lesions as found by Coolidge, Besic and Jacobs. A discussion of a recent theory of hydroxyapatite solubility of LaMer reveals calculation errors that vitiate his results. It is shown that logK ₁ (K ₁ is the ratio of the rate constants inq and can serve as a solubility equilibrium constant for hydroxyapatite) varies by only 1.2 units when calculated from three diverse sets of data. This variation is less than that reported by LaMer (when the errors of calculation in that work are corrected) and considerably less than the range of 11 among attempts to calculate a conventionalpK _sp , as summarized by Hodge. Literature list is found at the end of the last part, this issue, pp 462–464.     

Predicting kinetic constants of protein-protein interactions based on structural properties

Elucidating kinetic processes of protein–protein interactions (PPI) helps to understand how basic building blocks affect overall behavior of living systems. In this study, we used structure‐based properties to build predictive models for kinetic constants of PPI. A highly diverse PPI dataset, protein–protein kinetic interaction data and structures (PPKIDS), was built. PPKIDS contains 62 PPI with complex structures and kinetic constants measured experimentally. The influence of structural properties on kinetics of PPI was studied using 35 structure‐based features, describing different aspects of complex structures. Linear models for the prediction of kinetic constants were built by fitting with selected subsets of structure‐based features. The models gave correlation coefficients of 0.801, 0.732, and 0.770 for k_off, k_on, and K_d, respectively, in leave‐one‐out cross validations. The predictive models reported here use only protein complex structures as input and can be generally applied in PPI studies as well as systems biology modeling. Our study confirmed that different properties play different roles in the kinetic process of PPI. For example, k_on was affected by overall structural features of complexes, such as the composition of secondary structures, the change of translational and rotational entropy, and the electrostatic interaction; while k_off was determined by interfacial properties, such as number of contacted atom pairs per 100 Ų. This information provides useful hints for PPI design. Proteins 2010;79:720–734. © 2010 Wiley‐Liss, Inc.     

Application of a PEG precipitation method for solubility screening: A tool for developing high protein concentration formulations

Previous publications demonstrated that the extrapolated solubility by polyethylene glycol (PEG) precipitation method (Middaugh et al., J Biol Chem 1979; 254:367–370; Juckes, Biochim Biophys Acta 1971; 229:535–546; Foster et al., Biochim Biophys Acta 1973; 317:505; Mahadevan and Hall, AIChE J 1990; 36:1517–1528; Stevenson and Hageman, Pharm Res 1995; 12:1671–1676) has a strong correlation to experimentally measured solubility of proteins. Here, we explored the utility of extrapolated solubility as a method to compare multiple protein drug candidates when nonideality of a highly soluble protein prohibits accurate quantitative solubility prediction. To achieve high efficiency and reduce the amount of protein required, the method is miniaturized to microwell plate format for high‐throughput screening application. In this simplified version of the method, comparative solubility of proteins can be obtained without the need of concentration measurement of the supernatant following the precipitation step in the conventional method. The monoclonal antibodies with the lowest apparent solubilities determined by this method are the most difficult to be concentrated, indicating a good correlation between the prediction and empirical observations. This study also shows that the PEG precipitation method gives results for opalescence prediction that favorably compares to experimentally determined opalescence levels at high concentration. This approach may be useful in detecting proteins with potential solubility and opalescence problems prior to the time‐consuming and expensive development process of high concentration formulation.     

The role of subunit entropy in cooperative assembly. Nucleation of microtubules and other two-dimensional polymers.

The self-assembly and nucleation of two-dimensional polymers is described by a theory based on a model of rigid subunits and bonds and simple principles of thermodynamics. The key point in the theory is to separate as an explicit parameter the free energy, primarily attributed to the entropy of the free subunit, that is required to immobilize a subunit in the polymer. Quantitative relations for the association of a subunit forming a longitudinal bond, a lateral bone, or both together are obtained, which demonstrate the basis and magnitude of cooperativity. The same formalism leads to a quantitative estimate for th concentration of the small polymers that are important intermediates in nucleation. It is shown that, if the concentration of free subunits is below a certain "critical supersaturation," the concentration of some essential intermediates is too low to support any significant assembly and nucleation is blocked. If the subunit concentration is above the critical supersaturation, all of the small intermediates are sufficiently stable to form and grow spontaneously. The theory predicts a critical supersaturation of 3.5 to 7 (the ratio of subunit concentration to the equilibrium solubility) for parameters appropriate to assembly of the microtubule wall. Experimentally, nucleation and assembly of microtubules is obtained at somewhat lower concentrations, 1.5 to 3 times the equilibrium solubility. Special mechanisms that could stabilize small polymers and facilitate nucleation of microtubule assembly are suggested.     

Modeling of calcium-induced solubility profiles of casein for biotechnology: Influence of primary structure and posttranslational modification

Summary Molecular biology holds the promise of new tools for the food industry which include proteins with tailor-made functionality. Without a fundamental knowledge of the molecular bases of these properties, implementation will be strictly empirical. For example, the phenomena of salt-induced precipitation of proteins (salting-out) and their resolubilization (salting-in) has heretofore been discussed only qualitatively. A quantitative method, using Wyman's theory of thermodynamic linkage, has been developed and tested on the calcium-induced solubility profiles of the major milk proteins, the caseins. Salting-out was described by a salt-binding constant,k ₁, andn, the number of moles of salt bound; salting-in was described by the corresponding termsk ₂ andm. The magnitude of these parameters indicated involvement of protein phosphate groups in binding and precipitation, but enzymatic dephosphorylation showed significant increases ink ₁ andk ₂ indicating involvement of carboxylate groups as well. Studies on two genetic variants of _s1-casein indicated the importance of a hydrophobically stabilized intramolecular ion pair in the functionality of the protein. These studies have led to a fuller understanding of the molecular basis for the solubility behavior of caseins and have laid the groundwork for future computer simulation of food protein functionality.     

Mathematical modeling of precipitation and dissolution reactions in microbiological systems

We expand the biogeochemical model CCBATCH to include a precipitation/dissolution sub-model that contains kinetic and equilibrium options. This advancement extends CCBATCH's usefulness to situations in which microbial reactions cause or are affected by formation or dissolution of a solid phase. The kinetic option employs a rate expression that explicitly includes the intrinsic kinetics for reaction ormass-transport control, the differencefrom thermodynamic equilibrium, and the aqueous concentration of the rate-limiting metal or ligand. The equilibrium feature can be used alone, and it also serves as check that the kinetic rate never is too fast and ``overshoots' equilibrium. The features of the expanded CCBATCH are illustrated by an example in which the precipitation of Fe(OH)_{3 (s)} allows the biodegradation of citric acid, even though complexes are strong and not bioavailable. Precipitation releases citrate ligand, and biodegradation of the citrate increases the pH.     

Conditions leading to precipitation of ribulose bisphosphate carboxylase-oxygenase differ from those leading to enzyme activation

The relation between conditions leading to precipitation and/or activation of Ru-P₂ carboxylase have been explored in order to test the hypothesis that conformational changes leading to precipitation might be identical to those which are presumed to lead to enzyme activation. From the results of kinetic and solubility studies, we conclude that this hypothesis is not valid, since changes in solubility of Ru-P₂ carboxylase occur ten times as fast as changes in enzyme kinetics.     

Specific jarosite biomineralization by Purpureocillium lilacinum,an acidophilic fungi isolated from Río Tinto

Río Tinto (Huelva, southwestern Spain) is an extreme environment with a remarkably constant acidic pH and a high concentration of heavy metals, conditions generated by the metabolic activity of chemolithotrophic microorganisms thriving in the rich complex sulfides of the Iberian Pyrite Belt (IPB). Fungal strains isolated from the Tinto basin were characterized morphologically and phylogenetically. The strain identified as Purpureocillium lilacinum specifically induced the formation of a yellow‐ocher precipitate, identified as hydronium‐jarosite, an iron sulfate mineral which appears in abundance on the banks of Río Tinto. The biomineral was characterized by X‐ray diffraction (XRD) and its formation was observed with high‐resolution transmission electron microscopy (TEM) and scanning electron microscopy (SEM) coupled to energy‐dispersive X‐ray spectroscopy (EDX) microanalysis. Jarosite began to nucleate on the fungal cell wall, associated to the EPS, due to a local increase in the Fe³⁺/Fe²⁺ ratio which generated supersaturation. Its formation has been also observed in non‐viable cells, although with much less efficiency. The occurrence of P. lilacinum in an ecosystem with high concentrations of ferric iron and sulfates such as Río Tinto suggests that it could participate in the process of jarosite precipitation, helping to shape and control the geochemical properties of this environment.     

Characterization of maximal enzyme catalytic rates in central metabolism of Arabidopsis thaliana

Availability of plant‐specific enzyme kinetic data is scarce, limiting the predictive power of metabolic models and precluding identification of genetic factors of enzyme properties. Enzyme kinetic data are measured in vitro, often under non‐physiological conditions, and conclusions elicited from modeling warrant caution. Here we estimate maximal in vivo catalytic rates for 168 plant enzymes, including photosystems I and II, cytochrome‐b6f complex, ATP‐citrate synthase, sucrose‐phosphate synthase as well as enzymes from amino acid synthesis with previously undocumented enzyme kinetic data in BRENDA. The estimations are obtained by integrating condition‐specific quantitative proteomics data, maximal rates of selected enzymes, growth measurements from Arabidopsis thaliana rosette with and fluxes through canonical pathways in a constraint‐based model of leaf metabolism. In comparison to findings in Escherichia coli, we demonstrate weaker concordance between the plant‐specific in vitro and in vivo enzyme catalytic rates due to a low degree of enzyme saturation. This is supported by the finding that concentrations of nicotinamide adenine dinucleotide (phosphate), adenosine triphosphate and uridine triphosphate, calculated based on our maximal in vivo catalytic rates, and available quantitative metabolomics data are below reported values and, therefore, indicate undersaturation of respective enzymes. Our findings show that genome‐wide profiling of enzyme kinetic properties is feasible in plants, paving the way for understanding resource allocation.     

Copper(II)-DNA denaturation. I. Concentration dependence of melting temperature and terminal relaxation time

Both kinetic and equilibrium properties of DNA denaturation in the presence of copper(II) cation were studied by using optical techniques. Equilibrium properties of the reaction, measured in terms of T_m, the melting temperature, were shown to depend not on the overall but on the equilibrium concentrations of the species involved. Although T_m did increase with DNA concentration at low copper(II) concentrations, under similar conditions an increase in T_m with increasing copper(II) concentration was not observed. The kinetic properties of the reaction, characterized by the terminal relaxation time, were also found to depend on equilibrium concentrations of reactants. Application of standard methods of relaxation kinetics led to the proposal of a mechanism for copper(II)–DNA complex formation. That mechanism involves the rapid binding of two copper(II) ions to a reaction site on the polymer, followed by a slow, rate-limiting, first-order decay of the complex to the denatured state.     

Kinetic Resolution of Profens by Enantioselective Esterification Catalyzed by Candida antarctica and Candida rugosa Lipases

Profens (2‐arylpropionic acids) are known as one of the major nonsteroidal antiinflammatory drugs (NSAIDs) used in the treatment of inflammation associated with tissue injury. The inflammatory activity of profens is mainly due to their (S)‐enantiomer, whereas they are commercially available not only as pure enantiomers, but as racemates as well. There are several methods widely used in order to obtain enantiomerically pure compounds, however, the kinetic resolution with the application of lipases as biocatalysts may have an added advantage in the production of optically pure active pharmaceutical ingredients, such as milder reaction conditions, reduced energy requirements, and production costs. The aim of this study was to compare the results described in the literature in the case of the influence of reaction medium, alcohol moiety, and reaction temperature on the catalytic activity of lipases from Candida antarctica and Candida rugosa. Chirality 26:663–669, 2014. © 2014 Wiley Periodicals, Inc.     

Silica‐modified luminescent LaPO4:Eu@LaPO4@SiO2 core/shell nanorods: Synthesis,structural and luminescent properties

Monoclinic‐type tetragonal LaPO₄:Eu (core) and LaPO₄:Eu@LaPO₄ (core/shell) nanorods (NRs) were successfully prepared using a urea‐based co‐precipitation process under ambient conditions. An amorphous silica layer was coated around the luminescent core/shell NRs via the sol–gel process to improve their solubility and colloidal stability in aqueous and non‐aqueous media. The prepared nano‐products were systematically characterized by X‐ray diffraction pattern, transmission electron microscopy, energy dispersive X‐ray analysis, and FTIR, UV/Vis, and photoluminescence spectroscopy to examine their phase purity, crystal phase, surface chemistry, solubility and luminescence characteristics. The length and diameter of the nano‐products were in the range 80–120 nm and 10–15 nm, respectively. High solubility of the silica‐modified core/shell/Si NRs was found for the aqueous medium. The luminescent core NRs exhibited characteristic excitation and emission transitions in the visible region that were greatly affected by surface growth of insulating LaPO₄ and silica layers due to the multiphonon relaxation rate. Our luminescence spectral results clearly show a distinct difference in intensities for core, core/shell, and core/shell/Si NRs. Highly luminescent NRs with good solubility could be useful candidates for a variety of photonic‐based biomedical applications.     

Osmotic second virial cross‐coefficient measurements for binary combination of lysozyme,ovalbumin, and α‐amylase in salt solutions

Interactions measurement is a valuable tool to predict equilibrium phase separation of a desired protein in the presence of unwanted macromolecules. In this study, cross‐interactions were measured as the osmotic second virial cross‐coefficients (B₂₃) for the three binary protein systems involving lysozyme, ovalbumin, and α‐amylase in salt solutions (sodium chloride and ammonium sulfate). They were correlated with solubility for the binary protein mixtures. The cross‐interaction behavior at different salt concentrations was interpreted by either electrostatic or hydrophobic interaction forces. At low salt concentrations, the protein surface charge dominates cross‐interaction behavior as a function of pH. With added ovalbumin, the lysozyme solubility decreased linearly at low salt concentration in sodium chloride and increased at high salt concentration in ammonium sulfate. The B₂₃ value was found to be proportional to the slope of the lysozyme solubility against ovalbumin concentration and the correlation was explained by preferential interaction theory. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:1203–1211, 2013     

The preparation and kinetic properties of multiple forms of chicken brain acetylcholinesterase

A method for preparing various forms of acetylcholinesterase (A ChE) from chicken brain has been developed and they have been characterized in terms of kinetic parameters such as K_m, rate constant (k), turnover number (k_p), specificity constant (k_sp), V_max and half-life (t_1/2). The solubility experiments show that, there are two major forms of A ChE i.e. water-soluble and membrane-bound A ChE (MBA ChE). The MBA ChE shows several subforms, and on the basis of percentage activity only three MBA ChE forms have been selected for complete characterization by various kinetic parameters. It was found that these three forms of MBA ChE demonstrate significant differences in their kinetic properties.     

A regional impact assessment of climate and land-use change on alpine vegetation

Aim Assessing potential response of alpine plant species distribution to different future climatic and land‐use scenarios. Location Four mountain ranges totalling 150 km² in the north‐eastern Calcareous Alps of Austria. Methods Ordinal regression models of eighty‐five alpine plant species based on environmental constraints and land use determining their abundance. Site conditions are simulated spatially using a GIS, a Digital Terrain Model, meteorological station data and existing maps. Additionally, historical records were investigated to derive data on time spans since pastures were abandoned. This was then used to assess land‐use impacts on vegetation patterns in combination with climatic changes. Results A regionalized GCM scenario for 2050 (+ 0.65 °C, ?30 mm August precipitation) will only lead to local loss of potential habitat for alpine plant species. More profound changes (+ 2 °C, ?30 mm August precipitation; + 2 °C, ?60 mm August precipitation) however, will bring about a severe contraction of the alpine, non‐forest zone, because of range expansion of the treeline conifer Pinus mugo Turra and many alpine species will loose major parts of their habitat. Precipitation change significantly influences predicted future habitat patterns, mostly by enhancing the general trend. Maintenance of summer pastures facilitates the persistence of alpine plant species by providing refuges, but existing pastures are too small in the area to effectively prevent the regional extinction risk of alpine plant species. Main conclusions The results support earlier hypotheses that alpine plant species on mountain ranges with restricted habitat availability above the treeline will experience severe fragmentation and habitat loss, but only if the mean annual temperature increases by 2 °C or more. Even in temperate alpine regions it is important to consider precipitation in addition to temperature when climate impacts are to be assessed. The maintenance of large summer farms may contribute to preventing the expected loss of non‐forest habitats for alpine plant species. Conceptual and technical shortcomings of static equilibrium modelling limit the mechanistic understanding of the processes involved.