Determination of Binding Constants

This linearized form allows the straightforward determination of binding constants from the slopes of plots of effective mobilities vs. the term (ma — Meff )/ct. (Note The mobility of the free tBuCQN selector ma can be easily measured without additive and conversely, Meff of the selector is measured at various DNB-Leu concentrations.) The intercepts of such plots represent the effective mobilities of the complex, a figure... 

De Pauw, E. Determination of binding constants of oligonucleotide complexes with minor groove binders by electrospray ionization mass spectrometry. Comparison with fluorescence titration data. Adv. Mass Spectrom. 2001, 15, 795-796. 

Figure 4. Determination of binding constants of 2, 4 -dinitrophenyl 1-S-/ -cellobioside for CBH II by diafiltration (4°C).

KL Rundlett, DW Armstrong. Methods for the determination of binding constants by capillary electrophoresis. Electrophoresis 22 1419-1427, 2001. 

MHA Busch, JC Kraak, H Poppe. Principles and limitations of methods available for the determination of binding constants with affinity capillary electrophoresis. J. Chromatogr. A 777 329-353 (1997). 

Cyclodextrins are difficult to analyze because they show no appreciable UV absorbance and are often uncharged. Furthermore, they are not easy to label with visualizing agents because of their low reactivity. Therefore, cyclodextrins are often detected as inclusion complexes (45,46). A review dealing with the detection and separation of cyclodextrins as well as the determination of binding constants is given by Larsen and Zimmer-mann (47). 

A number of ACE assays dealing with the characterization of interactions between cyclodextrines as auxiliary substances and drugs have been presented. Other authors put more emphasis on the description of separation phenomenona by determination of binding constants. Model substances such as phenols are often used to examine the influence of ligand size and substitution as well as to evaluate the mathematical approaches for the calculation of binding constants. 

A complex overview of the current literature is given in Table 5. There the method used for the determination of binding constants is listed in ad-... 

Table 5 Papers Describing the Determination of Binding Constants Between CDs and Small Compounds... 

FA Gomez, LZ Avila, Y-H Chu, GM Whitesides. Determination of binding constants of ligands to proteins by affinity capillary electrophoresis compensation for electroosmotic flow. Anal Chem 66 1785-1791, 1994. 

E Szoko, J Gyimesi, L Barcza, K Magyar. Determination of binding constants and the influence of methanol on the separation of drug enantiomers in cyclodextrin-modified capillary electrophoresis. J Chromatogr A 745 181-187,... 

Alternative Capillary Electrophoresis Methods for Determination of Binding Constants... 

MT Bowser, DDY Chen. Monte Carlo simulation of error propagation in the determination of binding constants from rectangular hyperbolae. 2. Effect of the maximum response range. J Phys Chem 103 197-202, 1999. 

Dunn, B.M. and Chaiken, I.M., Quantitative affinity chromatography. Determination of binding constants by elution with competitive inhibitors, Proc. Natl. Acad. Set U.S.A., 71, 2382-2385, 1974. 

Pan J, Chen M, Warner W, He M, Dalton L, Hogen-Esch TE. Synthesis and self-assembly of diblock copolymers through hydrogen bonding. Semiquantitative determination of binding constants. Macromolecules 2000 33 7835-7841. 

Dufour, C. and Dangles, O., Flavonoid-serum albumin complexation determination of binding constants and binding sites by fluorescence spectroscopy, Biochim. Biophys. Acta, 1721, 164, 2005. 

Tran, C.D., and De Paoli Lacerda, S.H., Determination of binding constants of cyclodextrins in room-temperature ionic liquids by near-infrared spectrometry. Anal. Chem., 74, 5337-5341, 2002. 

Lastly, the use of micellar mobile phases allows a convenient means of studying micelle - solute interactions (i.e. determination of binding constants) (1,10 4,105) as well as determination of surfactant CMC values (from breaks in the log k gQ vs. log C, plots)... 

Equations are derived which take into account the formation of cyclodextrin to substrate complexes other than simple one to one host guest associations. An equation is also derived which describes the binding of a mono-protic species in which either its ionized or unionized form could bind to one or two cyclodextrin molecules. Because multiple binding constants are difficult to evaluate graphically, a non-linear least squares computer program is utilized. The approach works equally well for the determination of binding constants in micellar media. 

The binding constant between FITC-labeled bovine serum albumin and its monoclonal antibody was determined using ACE with LIF [14], Interaction of the enzyme cyclophilin with the immunosuppressive drug cyclosporine A was quantitatively assessed by CE with UV detection [15]. In the latter case the authors showed in particular how the difference between the sensitivity of the detector and KD of the studied system can severely hamper the correct determination of binding constants. 

The determination of binding constants, K, and inhibitor constants, Ki, for micelle-catalyzed reactions permits at least qualitative interpretations of the effect of substrate structure on the extent and nature of micellar complexation and allows a comparison of the magnitude of the binding constants for substrates in micellar systems with those in enzymatic systems. The hmited quantity of such data available at... 

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