Determination of Equilibrium Constants in Solution via ESI-MS

Electrospray mass spectrometry may appear a very attractive technique for the determination of the equilibrium constant of a reaction occurring in solution because of the soft nature of ESI, the simultaneous determination of the relative ion abundance of the reactants participating in the equilibrium, the high sensitivity, and the low sample consumption. 

The determinations of equilibrium constants using ESI-MS can be divided into two categories (a) equilibria ofsmall positive ions and ligands (such as crown ethers), so that the gas-phase ion creation involves lEM, and (b) equilibria of macro ions (such as proteins) and ligands (which may or may not be large organic molecules) so that gas-phase ion creation involves CRM. 

Consider the general reaction, Eq. (1.20a), where P is a ligand and S is a substrate, in a reaction that has reached equilibrium in the solution used. The equilibrium constant Kas is given by Eq. (1.20b) where [P], [S] and [PS] are the concentrations at equilibrium. 

Sampling the solution with ESI, the concentrations can be replaced with the ESI-MS observed peak intensities Ip, Is and Ips assuming that the individual sensitivity coefficients are equal to one. However this cannot be expected, and different strategies have been proposed to deal with this, see reference [54]. 

one can repeat the experiment at several, gradually increasing, concentrations of S and examine whether the association constant, evaluated with Eq. (1.20b), remains constant. This procedure is called the Titration Method and is generally used in ESI-MS determinations of the equilibrium constant. When the differences in molecular mass ofS, For PS are large, erroneous results may be obtained due to m/z-dependent transmission differences of the M S analyzer. While less of a problem for small metal (S)-ligand-(P) interactions, for proteins (P) of very large mass and substrate (S) of much smaller mass it is advantageous to use only the ratio /ps/lp. Zenobi and coworkers [55] have provided an equation for the determination of Kas in which only the ratio Ips/lp is used. 

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