Thermokinetic Method Research Articles

Gas phase basicity of X(CH 2) 3Y(X, Y = OH, NH 2) by the thermokinetic method

Gas phase basicity of three homologuous bidentate bases: 1,3-propanediamine, 1, 1,3-propanolamine, 2, and 1,3-propanediol, 3, has been determined by Fourier transform ion cyclotron resonance mass spectrometry. Both, the equilibrium and the thermokinetic methods were used and the results were compared with previous experimental and theoretical determinations. An excellent agreement is generally found between the gas phase basicities given by all the methods if corrections due to entropy variations are taken into account. In particular it is shown that a correction given by the term Δ G a ° = − ( T − 298 ) [ 〈 Δ p S 298 ° ( B i ) 〉 − Δ p S 298 ° ( M ) ] , where T is an “effective temperature” and Δ p S° the protonation entropy of the considered species, should be applied to the thermokinetic determination.

Microcanonical Modeling of the Thermokinetic Method

A microcanonical analysis of the thermokinetic method is performed using statistical rate calculations based on orbiting transition state theory in order to model a proton transfer process: MH(+) + B(i) --> M + B(i)H(+). The reaction efficiency is calculated as a function of the difference in zero point energy of reactants and products. Several models of reactions were investigated in order to simulate situations where the base of interest M exhibits loss of entropy upon protonation of up to approximately 40 J mol(-1) K(-1). It is shown that the standard thermokinetic method would predict correct 298 K gas phase basicities, GB(298)(M), even for polydentate molecules M, if experiments are conducted at this temperature. Proton affinity, PA(298)(M), and protonation entropy may be obtained by the thermokinetic method only in special circumstances such as, for example, experiments conducted at various temperatures.

Kinetic Studies on Na+/K+-ATPase and Inhibition of Na+/K+-ATPase by ATP

Na+/K+-ATPase (EC 3.6.1.3) is an important membrane-bound enzyme. In this paper, kinetic studies on Na+/K+-ATPase were carried out under mimetic physiological conditions. By using microcalorimeter, a thermokinetic method was employed for the first time. Compared with other methods, it provided accurate measurements of not only thermodynamic data (δrHm) but also the kinetic data (Km and Vmax). At 310.15 K and pH 7.4, the molar reaction enthalpy (δrHm) was measured as -40.514 ± 0.9 kJ mol−1. The Michaelis constant (Km) was determined to be 0.479 ± 0.020 mM and consistent with literature data. The reliability of the thermokinetic method was further confirmed by colorimetric studies. Furthermore, a simple and reliable kinetic procedure was presented for ascertaining the true substrate for Na+/K+-ATPase and determining the effect of free ATP. Results showed that the MgATP complex was the real substrate with a Km value of about 0.5 mM and free ATP was a competitive inhibitor with a Ki value of 0.253 mM.

Kinetic Studies on Na+/K+‐ATPase by Using Thermokinetic Method

AbstractNa+/K−‐ATPasc (EC 3.6.1.3) is an important membrane‐bound enzyme. By using microcalorimetry, the thermokinetic method was developed to kinetic studies on Na+/K+‐ATPase for the first time. Compared with other ones, the method provided accurate measurements of not only thermodynamic data but also the kinetic data. At 310.15 K and pH=7.4, the molar reaction enthalpy ΔrHm, was measured as (‐40.408k 1.9) kJ mol−1. The Michaelis constant Km was determined to be (0.479 0.020) 10−‐3 mol L−1 and consistent with literature figure which is about 0.5 10−‐3 moi L−1. The maximum velocity Vmax obtained was (0.6810.026) μmol Pi min−1 mg protein−1. All of the data have good repeatability and self‐consistency. The reliability of thermokinetic method was verified by the experimental results and further confirmed by colorimetric studies. Moreover. the effect of enzyme pre‐dilution on its activities was also investigated.

Microcalorimetric Studies on the Product Inhibition of Hydrolysis of L-arginine Catalyzed by Bovine Liver Arginase

A new thermokinetic reduced extent method for the product inhibition of single substrate enzyme-catalyzed reactions is proposed and compared with the traditional initial rate method in this paper. The arginase-catalyzed hydrolysis of L-arginine to L-ornithine and urea was studied at 37°C in 40 mM sodium barbiturate-HCl buffer solution (pH=9.4). Michaelis constant (K m ) for arginine and maximum velocity (V m ) of the reaction were determined by initial method and thermokinetic method. The activation of exogenous manganese to this reaction was also studied. The product inhibition constant (K p ), which cannot be obtained directly from the initial rate method, was determined by thermokinetic without adding L-ornithine to the reaction system. When the concentration of Mn 2+ in cell is 0.1 mM, the enzyme gets its full activity. Incubation arginase with appropriate concentration of Mn 2+ resulted in increased V max and a higher sensitivity of the enzyme to product with no change in the K m for arginine. We suggest that the exogenous manganese ions in solution have just recovered the activity of arginase, which was lost in dissolving and dilution, but no effect on the mechanism of the reaction.

Characteristic parameter method for studying chemical kinetics in calorimeter

A novel thermokinetic research method for determination of rate constants of simple-order reaction in batch conduction calorimeter under isothermal condition, the characteristic parameter method, is proposed in this paper. Only needing the characteristic time parameter tm obtained from the measured thermoanalytical curve, the kinetic parameters of reactions studied can be calculated conveniently with this method. The saponifications of ethyl propionate and ethyl acetate in aqueous ethanol solvent, the polymerization of acrylamide in aqueous solution, the ring opening reaction of epichlorohydrin with hydrobromic acid have been studied. The experimental results indicate that the characteristic parameter method for simple-order reaction is correct.

Thermokinetic studies on the activation of bovine liver arginase by manganese ions

The activation of bovine liver arginase, which catalyzes the hydrolysis of l-arginine to l-ornithine and urea, by manganese ions was studied by thermokinetic methods at 37 °C in 40 mM sodium barbiturate–HCl buffer solution (pH 9.4). Full activation of arginase, by incubation with 0.1 mM Mn 2+, resulted in increased of V max, and a higher sensitivity of the enzyme to product and l-lysine inhibition, with no change in the K m for arginine. Upon addition of 0.1 mM Mn 2+ to the reaction, the inhibitory constants of product ( K P) and l-lysine ( K I) decreased from 1.18 to 0.70 mM and from 5.60 to 3.10 mM, respectively. We suggest that the exogenous manganese ions in reaction recovered the activity of arginase, which was lost in dissolving and dilution, without effecting on the mechanism of the reaction.

The kinetic energy dependence of association reactions. A new thermokinetic method for large systems

The reactions of bare alkali metal ions (M+=Li+, Na+, or K+) with dimethoxyethane (CH3OCH2CH2OCH3, DXE) are studied using guided ion beam tandem mass spectrometry. The bimolecular reaction forms an associative M+(DXE) complex that is long-lived and dissociates back to the reactants. The kinetic energy dependences of the cross sections for formation of the complexes are interpreted with several different models (including rigorous phase space theory) that assume that the complex lifetimes are limited by dissociation over a loose, orbiting transition state. After accounting for the effects of multiple ion–molecule collisions, internal energy of the reactant ions, Doppler broadening, and dissociation lifetimes, the analyses yield 0 K bond energies as the only adjustable parameter. These values are compared with bond energies obtained from previous collision-induced dissociation (CID) studies of the M+(DXE) complexes and found to be self-consistent for all models studied. Association and CID form the same energized M+(DXE) complex in two distinct ways, such that a comparison of these results allows an assessment of the models used to interpret CID thresholds and test the limits of statistical theories such as RRKM and phase space theory.

Gas-phase basicity of glycine, alanine, proline, serine, lysine, histidine and some of their peptides by the thermokinetic method.

The thermokinetic method is applied to a set of six amino acids (glycine, alanine, proline, serine, lysine, histidine) and 30 of their di- and tri-peptides for which experimental proton transfer rate constants were available. The comparison between the presently determined gas-phase basicities, GBs, of the amino acids with values obtained from equilibrium constant determination is generally good (a mean deviation of appoximately 3 kJ mol(-1) is observed). Derived proton affinities values are discussed. The gas-phase basicities of peptides provided by the present study correct several previously estimated values thus offering a more firm basis for structural discussion. Composite reaction efficiency curves indicate the existence, for several peptides, of at least two non-interconverting populations of protonated forms.

The gas-phase basicity and proton affinity of 1,3,5-cycloheptatriene--energetics, structure and interconversion of dihydrotropylium ions.

The hitherto unknown gas-phase basicity and proton affinity of 1,3,5-cycloheptatriene (CHT) have been determined by Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry. Several independent techniques were used in order to exclude ambiguities due to proton-induced isomerisation of the conjugate cyclic C(7)H(9)(+) ions, [CHT + H](+). The gas-phase basicity obtained by the thermokinetic method, GB(CHT) = 799 +/- 4 kJ mol(-1), was found to be identical, within the limits of experimental error, with the values measured by the equilibrium method starting with protonated reference bases, and with the values resulting from the measurements of the individual forward and reverse rate constants, when corrections were made for the isomerised fraction of the C(7)H(9)(+) population. The experimentally determined gas-phase basicity leads to the proton affinity of cycloheptatriene, PA(CHT) = 833 +/- 4 kJ mol(-1), and the heat of formation of the cyclo-C(7)H(9)(+) ion, deltaH(f)(0)([CHT + H](+)) = 884 +/- 4 kJ mol(-1). Ab initio calculations are in agreement with these experimental values if the 1,2-dihydrotropylium tautomer, [CHT + H((1))](+), generated by protonation of CHT at C-1, is assumed to be the conjugate acid, resulting in PA(CHT) = 825 +/- 2 kJ mol(-1) and deltaH(f)(0)(300)([CHT + H((1))](+)) = 892 +/- 2 kJ mol(-1). However, the calculations indicate that protonation of cycloheptatriene at C-2 gives rise to transannular C-C bond formation, generating protonated norcaradiene [NCD + H](+), a valence tautomer being 19 kJ mol(-1) more stable than [CHT + H((1))](+). The 1,4-dihydrotropylium ion, [CHT + H((3))](+), generated by protonation of CHT at C-3, is 17 kJ mol(-1) less stable than [CHT + H((1))](+). The bicyclic isomer [NCD + H](+) is separated by relatively high barriers, 70 and 66 kJ mol(-1) from the monocyclic isomers, [CHT + H((1))](+) and [CHT + H((3))](+), respectively. Therefore, the initially formed 1,2-dihydrotropylium ion [CHT + H((1))](+) does not rearrange to the bicyclic isomer [NCD + H](+) under mild protonation conditions.

Cyclopentenyl cation: its thermochemistry and its characterized formation from C 6H 10[rad]+ species

The formation of cyclopentenyl cation from (i) dissociation of ionized cyclohexene, and, (ii) ion–molecule reaction between ionized 1,3-butadiene and ethylene is confirmed by means of the corresponding thermochemistry. Measurement of the Gibbs free energy and enthalpy for deprotonation of cyclopentenyl cation is provided by Fourier transform ion cyclotron resonance experiments and use of the thermokinetic method. The values (GB=795.0±1.2 and PA=824.9±1.1 kJ/ mol , respectively) allow the determination of the heat of formation of the cyclopentenyl cation to be 840.6±4.1 kJ/ mol in good agreement with existing data.

Thermodynamics and kinetics of the cleavage of DNA catalyzed by bleomycin A5.

Microcalorimetry and UV-vis spectroscopy were used to conduct thermodynamic and kinetic investigations of the scission of calf thymus DNA catalyzed by bleomycin A5 (BLM-A5) in the presence of ferrous ion and oxygen. The molar reaction enthalpy for the cleavage, the Michaelis-Menten constant for calf thymus DNA and the turnover number of BLM-A5 were calculated by a novel thermokinetic method for an enzyme-catalyzed reaction to be -577 +/- 19 kJ.mol-1, 20.4 +/- 3.8 microm and 2.28 +/- 0.49 x 10-2 s-1, respectively, at 37.0 degrees C. This DNA cleavage was a largely exothermic reaction. The catalytic efficiency of BLM-A5 is of the same order of magnitude as that of lysozyme but several orders of magnitude lower than those of TaqI restriction endonuclease, NaeI endonuclease and BamHI endonuclease. By comparing the molar enthalpy change for the cleavage of calf thymus DNA induced by BLM-A5 with those for the scission of calf thymus DNA mediated by adriamycin and by (1,10-phenanthroline)-copper, it was found that BLM-A5 possessed the highest DNA cleavage efficiency among these DNA-damaging agents. These results suggest that BLM-A5 is not as efficient as a DNA-cleaving enzyme although the cleavage of DNA by BLM-A5 follows Michaelis-Menten kinetics. Binding of BLM-A5 to calf thymus DNA is driven by a favorable entropy increase with a less favorable enthalpy decrease, in line with a partial intercalation mode involved in BLM-catalyzed breakage of DNA.

Bleomycin A5 is analogous in mechanism to a DNA-cleaving enzyme: a microcalorimetric study

Microcalorimetry, UV–VIS spectroscopy and HPLC were used to conduct thermodynamic and kinetic investigations of the scission of calf thymus DNA catalyzed by bleomycin A5 (BLM-A5) in the presence of ferrous ion and oxygen. The molar reaction enthalpy for the cleavage, the Michaelis constant for calf thymus DNA and the turnover number of BLM-A5 were calculated by a novel thermokinetic method for enzyme-catalyzed reactions to be −577±19kJmol−1, (2.04±0.38)×10−5moldm−3 and (2.28±0.49)×10−2s−1, respectively, at 37.00°C. This DNA cleavage was a fast and exothermic reaction. The catalytic efficiencies of BLM-A5 and of some DNA-cleaving enzymes, such as TaqI restriction endonuclease and NaeI endonuclease, are of the same order of magnitude. By comparing the molar enthalpy change for the cleavage of calf thymus DNA induced by BLM-A5 with those for the scission of calf thymus DNA mediated by adriamycin and by (1,10-phenanthroline)-copper, it was found that BLM-A5 possessed the highest DNA cleavage efficiency among these DNA-damaging agents. The UV–VIS spectral experiments showed that BLM-A5 itself remained unchanged chemically after the cleavage. The products of this scission, including four kinds of free nucleic acid bases, were separated and detected by HPLC. These results strongly suggest that BLM-A5 is analogous in mechanism to a DNA-cleaving enzyme, although it is much smaller than such type of enzymes. A possible mechanism for the cleavage of DNA catalyzed by BLM was suggested in this paper.

Characteristic parameter method for studying kinetics of opposing reactions in calorimeter

According to the theoretical basis of thermokinetics, the integral and differential thermokinetic equations of opposing reactions have been derived, and a novel thermokinetic research method, the characteristic parameter method for opposing reactions which taking place in a batch conduction calorimeter under isothermal condition, has been proposed in this paper. Only needing the characteristic thermoanalytical data corresponding to tm and 2tm from the same curve, the rate constants of forward and backward reactions and equilibrium constant can be calculated simultaneously with this method. In order to test the validity of this method, the proton-transfer reactions of nitroethane with ammonia at 15 and 25°C, and with trihydroxymethyl aminomethane (Tris) at 15 and 30°C have been studied, respectively. The results of rate constants and equilibrium constants calculated with this method are in agreement with those in the literature. Therefore, the characteristic parameter method for opposing reaction is believed to be correct.

Determination of ionization energies by the thermokinetic method

Charge transfer reactions of the type M·++B → B·++M have been studied in order to test the applicability of the “thermokinetic” method to the determination of ionization energies (IEs). Three examples are presented with M=benzene, cyclopentanone, and vinyl alcohol. The correlation observed between the bimolecular rate constants and IE(B) allows the determination of IE(M) with a mean deviation of ∼0.02 eV with respect to spectroscopic values. One other advantage of the method is to provide ionization energies of unstable neutrals. The role of various corrective terms, and of possible geometry changes during charge transfer reactions are discussed.

Time-parameter method for studying kinetics of consecutive first-order reactions in calorimeter

In order to enrich the thermokinetic research methods and enlarge the applicable range of the thermokinetic time-parameter method, the integral and differential thermokinetic equations of consecutive first-order reaction have been deduced, and the mathematical models of the time-parameter method for consecutive first-order reactions have been proposed in this paper. The rate constants of two steps can be calculated from the same thermoanalytical curve measured in a batch conduction calorimeter simultaneously with this method. The thermokinetics of saponifications of diester in aqueous ethanol solvent has been studied. The experimental results indicate that the time-parameter method for the consecutive first-order reaction is correct.

Microcalorimetric studies on catalase reaction and inhibition of catalase by cyanide ion

As Chance et al. [1–3] proposed, the decomposition of hydrogen peroxide catalyzed by catalase is an overall first-order reaction. In this paper, we have studied this enzyme-catalyzed reaction with a thermokinetic method. The rate constant and the molar reaction enthalpy of this reaction have been measured. At 310.15 K and pH=8.2, k cat=1.75×10 6 l mol −1 s −1, Δ r H m=88.99 kJ mol −1. Furthermore, we have studied the competitive inhibition of catalase by cyanide ion and reported some correlated parameters.

TG-DTA studies on wood treated with flame-retardants

wood untreated and treated with flame-retardants were analyzed using the TG-DTA simultaneous method. Results showed that upon lowering the pyrolysis temperature, wood weight loss was reduced and char yield increased in the charring phase of the pyrolysis. The TG curves were further analyzed by the thermokinetic method to obtain thermokinetic parameters of the different phases during the wood pyrolysis and to derive a thermokinetic Equation. It was found that the activation energy in the drying phase was reduced and the activation energy in the charring or calcining phase varied for the different flame-retardants used.

Gas-phase basicities of the isomeric dihydroxybenzoic acids and gas-phase acidities of their radical cations

The thermochemical acid/base properties of the six dihydroxybenzoic acids ( x, y-DHB) as prototypical matrices used in matrix-assisted laser desorption/ionization (MALDI) have been investigated. The ground-state gas-phase basicities (GB) of the six DHB isomers and the gas-phase acidities (Δ G acid) of the corresponding radical cations ([ x, y-DHB] ·+) have been determined by Fourier-transform ion cyclotron resonance mass spectrometry employing the thermokinetic method. The gas-phase basicities vary from 814 kJ mol −1 for the least basic isomer, 3,5-DHB, to 831 kJ mol −1 for the most basic isomer, 2,4-DHB. The obtained gas-phase acidities of the corresponding radical cations vary from 815 kJ mol −1 for the most acidic species, 3,4-DHB, to 858 kJ mol −1 for the least acidic one, 2,5-DHB. The results indicate that ground-state proton transfer from the matrix radical cations to the analyte may play a role in the ionization process of MALDI, whereas proton transfer from protonated matrix molecules can be excluded.

Gas-Phase Basicity of Formaldehyde by the Thermokinetic Method

A series of proton transfer reactions monitored in a Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometer allows the determination of the gas-phase basicity ( GB) and the proton affinity ( PA) of formaldehyde. The values determined by the thermokinetic method, GB(CH2O) = 681.5 ± 0.7 kJ mol−1 and PA(CH2O) = 711.5 ± 2.1 kJ mol−1 are in excellent agreement with data originating from proton transfer equilibrium constant determinations or from G2 calculations.