mass spectrometry; noncovalent complexes; binding constants; oligonucleotides; cyclodextrins; DNA
Abstract :
[en] A method for determining the equilibrium association constant of a complexation reaction A + B <=> AB by electrospray ionization mass spectrometry is described. The method consists in measuring the relative intensities of the peaks corresponding to A and to AB in equimolar A–B solutions at different concentrations C0. The results are fitted by a non-linear least-squares procedure, with the two variable parameters being the equilibrium association constant Ka and a factor R, defined by I(AB)/I.A/ = R× [AB]/[A]. The factor R is the ratio between the response factors of AB and A, and corrects
for the relative electrospray responses of the complex and the free substrate A, mass discrimination of instrumental origin and/or moderate in-source dissociation. The method is illustrated with the following two systems: complexes between a double-stranded 12-base pair oligonucleotide and minor groove binders, and cyclodextrin complexeswith a,!-dicarboxylic acids. For the oligonucleotide complexes, it is found that the response of the complex is not dramatically different to the response of the free oligonucleotide duplex, as the double helix conformation is disturbed by the drug only to a minor extent. In the case of cyclodextrin complexes, these complexes were found to have a much higher response than free cyclodextrin. This may be due to the fact that cyclodextrin is neutral in solution, whereas the complex is charged, but it can also stem from the fact that a significant proportion of the complex is in a non-inclusion geometry. The present method requires the exact determination of the concentrations of the reactants and is applicable to 1 : 1 complexes.
Research center :
CART - Centre Interfacultaire d'Analyse des Résidus en Traces - ULiège
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