A detailed examination of boronic acid-carbohydrate complexation using alizarin red S. as a general optical reporter

Boronic acids and compounds containing diol moieties bind with high affinity through reversible ester formation. The conditions that foster tight binding between the diol and the boronic acid have heretofore not been well understood. Due to the multiple ionic states of both the boronic acid and its ester, the equilibrium constants reported in the literature have not always been strictly defined, and therefore, there is a lack of comparability among the reported values. To address these issues, we have developed a method for examining boronate ester stability using a three component competitive system containing the fluorescent reporter Alizarin Red S. Alizarin Red S. displays a dramatic change in fluorescence intensity and absorbance spectrum in response to the binding of a boronic acid and can be used as a general reporter for studying carbohydrate-boronic acid interactions, both quantitatively and qualitatively. We have used this system to determine the association constants of a series of diols, and as a basis from which to derive a number of relationships that correlate the various equilibrium constants in the literature. This study has also allowed us (1) to understand the trends of pH versus boronate ester stability, and (2) to develop a predictive methodology that examines the relative stabilities of boronate esters formed from different carbohydrates.