Dissociation curve analysis

Goodford PI, St-Louis J, Wootton R. A quantitative analysis of the effects of 2,3-diphosphoglycerate, adenosine triphosphate and inositol hexaphosphate on the oxygen dissociation curve of human haemoglobin. J Physiol 1978 283 397. 

Figure 3 Dependence of the equilibrium concentration [A] of the analyte on the initial concentration [B0] (indicated by (B) in the text) of ligand added to the reaction mixture before CZE analysis. The curves are calculated for different values of the dissociation constants (between 1CT5- and 103-fold of the initial concentration of A). Reaction between analyte and ligand is according to A + B = AB. The initial concentration (A) is kept constant, and increasing initial concentrations (B) are added to the reaction mixture. All concentrations are given in arbitrary units (including the dimension of KD). (Reprinted from Ref. 22.)...

At present researchers and clinicians interested in complete blood gas analysis must resort to different sets of instruments and techniques for each of the three types of measurement. This chapter describes a membrane permeation cell connected to a mass spectrometer that can measure blood gas partial pressures, contents, and the position of the 02 dissociation curve. 

Dissociation Curve Analysis. Torrance and Lenfant (29) grouped the determination of the dissociation curve into three different methodologies. The first measures the 02 content or saturation over a set of known po2 s. The second approach measures the blood p02 using samples at known saturation. The most frequent determination is at the 50% saturation level described by Edwards and Martin (30). 

Though the most direct application of the analysis system is determining gas contents, the construction of the 02 dissociation curve requires computation of Po2s at several positions. This can introduce errors of the type mentioned. We feel that the potential of rapidly obtaining a dissociation curve for clinical use will more than balance these discrepancies. 

For research applications, where maximal accuracy is desirable even at the expense of time and effort, two maneuvers are possible. The first involves derivation of the initial dissociation curve using the standard calibration curve. A second computation follows, using the derived curve for calibration. This process is repeated until the derived and calibrating dissociation curves match each other. This method requires considerable calculations, but it is theoretically valid even for abnormal hemoglobins. The other technique for accurate dissociation curve analysis is simply a variation on the traditional method of carefully equilibrating aliquots... 

Maruyama, K. Takeyama, H. Nemoto, E. Tanaka, T. Yoda, K Matsunaga, T. (2004), Single nucleotide polymorphism detection in aldehyde dehydrogenase 2 (ALDH2) gene using bacterial magnetic particles based on dissociation curve analysis. 

This analysis was performed separately for Cy3-cAMP bound to the anterior pseudopods and the posterior tails of the Dictyostelium cells undergoing chemotaxis, revealing that Cy3-cAMP receptor complexes on anterior pseudopods dissociated about three times faster than those on posterior tails (Fig. 13). The dissociation curves were fitted to a sum of two exponential functions, indicating the presence of at least two receptors that adopt different kinetic states. Further characterization of this difference in the receptor states between anterior and posterior region suggests that the difference in the receptor states... 

The initial goal of the kinetic analysis is to express k as a function of [H ], pH-independent rate constants, and appropriate acid-base dissociation constants. Then numerical estimates of these constants are obtained. The theoretical pH-rate profile can now be calculated and compared with the experimental curve. A quantitative agreement indicates that the proposed rate equation is consistent with experiment. It is advisable to use other information (such as independently measured dissociation constants) to support the kinetic analysis. 

A frequently encountered pH-rate profile exhibits a bell-like shape or hump, with two inflection points. This graphical feature is essentially two sigmoid curves back-to-back. By analogy with the earlier analysis of the sigmoid pH-rate curve, where the shape was ascribed to an acid-base equilibrium of the substrate, we find that the bell-shaped curve can usually be accounted for in terms of two acid-base dissociations of the substrate. The substrate can be regarded, for this analysis, as a dibasic acid H2S, where the charge type is irrelevant we take the neutral molecule as an example. The acid dissociation constants are... 

It was concluded [734] from visual inspection and chemical analysis of partially decomposed dolomite, that reaction was initiated at the outer surfaces of the crystallites and the interface established advanced thereafter into the bulk. The deceleratory a—time curves obeyed the contracting volume equation [eqn. (7), n = 3] and the values of E determined were between 206 and 232 kJ mole-1. These values of E were generally greater than those reported for other studies ( 190 kJ mole-1) which are in the range mentioned [121] for CaC03 dissociation and slightly larger than the enthalpy of that reaction. On exposure of the residue from vacuum decomposition of dolomite to C02, the gas uptake at 1070 K was... 

Figure 4. Continuous analysis by fluorescence polarization of FLPEP binding and dissociation to permeabilized neutrophils. Each curve represents the average of two determinations. The descending curves represent the effect of the addition of receptor antagonist after FLPEP had bound to the receptors. Upper panel, in the absence of GTPyS, receptor antagonist added at t = 60 s lower panel, in the presence of GTPyS, antagonist added at t =...

Provided that a value for KL is available, it is possible to use this equation to obtain a value for Kh the dissociation equilibrium constant for the inhibitor, by nonlinear least-squares analysis of the displacement curve. Alternatively, K can be calculated from the IC50, which may be obtained by simple interpolation by eye from a Hill plot or by fitting a curve to an equation of the type ... 

This technique was employed to study the binding dynamics of Pyronine Y (31) and B (32) with /)-CD/ s The theoretical background for this particular system has been discussed with the description of the technique above. Separate analysis of the individual correlation curves obtained was difficult since the diffusion time for the complex could not be determined directly because, even at the highest concentration of CD employed, about 20% of the guest molecules were still free in solution. The curves were therefore analyzed using global analysis to obtain the dissociation rate constant for the 1 1 complex (Table 12). The association rate constant was then calculated from the definition of the equilibrium constant. 

For the determination of the dissociation constant in the excited state, several methods have been used the Forster cycle,(109 m) the fluorescence titration curve/113 the triplet-triplet absorbance titration curve,014 but all involve the assumption that the acid-base equilibrium may be established during the lifetime of the excited state, which is by no means a common occurrence. A dynamic analysis using nanosecond or picosecond time-resolved spectroscopy is therefore often needed to obtain the correct pK a values.1(n5)... 

An indication has been obtained that the opening of the salicylate hydrogen bond may become partially rate limiting in proton transfer (33) from substituted salicylate ions to hydroxide ions and buffer species in 50% (v/v) MejSO-HjO (Hibbert and Spiers, 1989a). Temperature-jump measurements of the equilibration between the salicylate ion and its dissociated species lead to curved plots of against buffer concentration and against hydroxide-ion concentration. Analysis of the results in terms of the mechanism in (33) gave the approximate values ki = 5x 10 s" and A, = 3 X 10 s ... 

The equilibrium dialysis experiment revealed that histidine-substituted salicylamide was selected as an RNA ligand. Subsequent binding analysis by UV titrations and Job plot revealed the histidine-substituted salicylamide Cu + complex bound the target RNA hairpin with an apparent dissociation constant of 150 nM. This binding constant likely reflects more complex binding processes than a simple 1 1 interaction, as the observed binding curve saturates well below the concentration of the histidine-substituted salicylamide, and thus the actual affinity of the complex for targeted RNA is probably lower. Importantly, however, titrations with the... 

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