Equilibrium dialysis method

Protein binding parameters B, and Kh as depicted in Eq. (44), are obtained from in vitro experiments utilizing filtration, centrifugation, and dynamic dialysis or equilibrium dialysis methods. These techniques have been reviewed elsewhere [54,55],... 

Kariv, I., Cao, H. and Oldenburg, KR. (2001) Development of a high throughput equilibrium dialysis method. Journal of Pharmaceutical Sciences, 90, 580-587. 

In spite of much information available for the interactions of various metal ions with small oxoanions of phosphorus, relatively little information has been obtained for the complex formation of long-chain polyphosphate ion. This may be due to the fact that the conventional methods useful for the study of the complex formation of a relatively small ligand are not always applicable to the polyanion complex formation system. Since a gel chromatographic method based on the same principle as the equilibrium dialysis method has been proved to be applicable in the field of inorganic complex chemistry (1), this method has been applied to the study of the binding of long-chain polyphosphate ions to magnesium ion. 

FIGURE 3.31 Equilibrium dialysis method for analysis of drug-protein binding. 

Kariv I, Cao H, Oldenburg KR (2000) Development of a High Throughput Equilibrium Dialysis Method. J Pharma Sci 90 580-587... 

Other methods to determine the interactions between aroma compounds and wine matrix components do not involve gas phase measurements. For example, the equilibrium dialysis method has been applied for determining interactions between yeast macromolecules and some wine aroma compounds (Lubbers et al. 1994a) and more recently to study the interaction of aroma compounds and catequins in aqueous solution (Jung and Ebeler 2003). While this method can be set up in different ways, a simple approach is to fill a dialysis cell (two chambers separated by a semiper-meable membrane) with an aromatized liquid. A non-volatile component of wine can be added to one chamber of the cell and then the system allowed to come to equilibrium. If the added non-volatile component binds the aroma compound, the other chamber will be depleted by this binding. Quantification of this change in concentration permits calculating the quantity that is bound to the added substrate. 

Equilibrium Dialysis Method. The equilibrium dialysis method is based on the diffusion of the volatile compound through a semi-permeable membrane placed between two compartments containing the model wine and macromolecules (11). In the experiment, ImL solution of macromolecule in the model wine was placed on one side of the membrane (compartment 1) and 1 mL of the model wine containing a known amount of the volatile compound on the other side (compartment 2). The system was shaken at 30 °C for 12 h to reach equilibrium of the free ligand (volatile compound) between the two compartments of the cell. At equilibrium, the concentration of the volatile compound was determined by gas chromatography. The difference in concentration of the volatile compound between the two compartments represents the amount of the volatile compound bound to the macromolecule. 

Figure 1. Percentage of binding with F extract and the fractionsN-Fl and R-Fl (from affinity chromatography on Con-A) at 10 g/L in model wine by the equilibrium dialysis method.

Effect of Ethanol on Conformational state of protein. To understand the effect of ethanol and pH in flavour-protein interactions the binding of y-decalactone to bovine serum albumin was investigated using the equilibrium dialysis method (20). Without ethanol, a decrease in pH (from 5.3 to3.5) reduces by one-half the y-decalactone binding onto protein. In the presence of ethanol, changing pH do not have any appreciable effect (Table VI). 

Several experimental methods based on diflferent principles have been proposed for the study of polyion-small ion-binding equilibria they include an electrochemical method, mainly through application of ion-specific electrodes [40-44], an equilibrium dialysis method [45], a dye method [46-51], and an ion-exchange distribution method [52-56]. In any case, the free ion concentration in the system under investigation is determined. They permits the evaluation of 0 with Eq. (4). The binding quotient, Ka. used to express the overall (apparent) equilibrium is then given by Eq. (5). 

Figure 14.10. Elution profile of RNase as a function of free [5 -TMP], using an affinity column with immobilized 5 -TMP. The concentrations of 5 -TMP in the mobile phase were 5.0 x 10 4Af, 4.0 x 10 4Af, 3.0 x 10 4Af,2.0 x 10 4M, 1.0 x 10 4M, and 7.5 x 10 5M for the earliest to the latest eluted fractions, respectively. The inset shows the plot according to Eq. 14.24 that was used to determine the association constant for the RNase-5 -TMP binding reaction.11 [Reprinted, with permission, from B. M. Dunn and I. M. Chaiken, Biochemistry 14 (No. 11), 1975, 2343-2349. Evaluation of Quantitative Affinity Chromatography by Comparison with Kinetic and Equilibrium Dialysis Methods for the Analysis of Nucleotide Binding to Staphylococcal Nuclease . 1975 by American Chemical Society.]...

The need for prolonged dialysis inherent in equilibrium dialysis methods has been circumvented by the development of a method in which a flow-dialysis, or rate of dialysis, procedure is employed (56). This method employs a dialysis cell with an upper chamber, containing the... 

Explain equilibrium dialysis methods for measuring binding equilibrium... 

Equilibrium dialysis methods often involve the use of radioactively labelled ligands, so that very small concentrations and very tight binding can be measured directly. 

Equilibrium dialysis methods can be used to measure binding interactions directly. 

The structural and catalytic zinc atoms have also been replaced by cadmium-109 by equilibrium dialysis methods similar to that for cobalt, again either replacing just the structure zinc atoms giving [(LADH)Cd2Zn2], or both types of zinc giving [(LADH)Cd2Cd2]. The enzymic activity of both species was similar to the native enzyme. The cadmium-substituted enzyme is similar in all other respects to the cobalt substituted enzyme, i.e. mode of inhibition, UV spectra. The CdNMR spectrum has been observed for the Cd fully substituted enzyme and, as expected, shows two resonances corresponding to the two types of cadmium atoms in the enzyme. ... 

The classical methods of studying protein binding of drugs include equilibrium dialysis and ultrafiltration. The latter may provide quick measurements but is not necessarily as accurate as the equilibrium dialysis method. Detailed discussions on this may be found in textbooks listed in Suggested Readings. 

Binding ratios of SDS (BHD Ltd.) and CPC (Tokyo Kasei Ltd.) to HPC (M.W. =11 -I5xl0 Tokyo Kasei Ltd.) were determined at 30 °C by an equilibrium dialysis method. Cloud point of an HPC solution was observed by the naked eye. Amounts of p-dimethylaminoazobenzene (Nakarai Ltd.) solubilized by the surfactant micelle and the surfactant-polymer complex were determined at 30 °C by colorimetry at 1 = 418 nm. Viscosity was measured by an Ubbelohde-type capillary viscometer, a cone-plate rotary viscometer, or a Brookfield-type rotary viscometer at 25 or 30 °C. Mean diameter of secondary particles of kaolinite in its dilute suspension (1 g/dl) was estimated by a Coulter counter (type TA-II) in 154 mmol/dm NaCl at room temperature. 

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