Molecular crowding effect on ion-pair extraction of picric acid and alkylammonium ion.

Macromolecular crowding, a highly concentrated environment of macromolecules, can alter chemical equilibria and reaction kinetics. Although molecular crowding has been recognized as a useful strategy for enhancing reaction efficiency, how it affects individual elementary equilibria within a coupled chemical process remains less understood. In this study, we investigated the effects of macromolecular crowding on the ion-pair extraction of picric acid with tetraalkylammonium ions using dextran with molecular weights of 40 kDa (Dex40k) and 200 kDa (Dex200k) as crowding agents. The extraction constant (Kex), distribution coefficient (KD), and association constant (KA) were determined using spectrophotometry. For the Dex40k system, Kex and KA increased with dextran concentration, whereas KD decreased. These changes were analyzed using a thermodynamic model based on the excluded volume and osmotic pressure effects, which are considered the principal origins of molecular crowding in cells. The analysis revealed that the increase in Kex was primarily driven by the excluded volume effect, whereas the decrease in KD was primarily governed by the osmotic pressure effect. In contrast, both effects contributed to the enhancement of the KA. Furthermore, experiments using Dex200k, which forms a polymer-network structure, suggested that the excluded volume effects may remain involved even in polymer-network environments. These results demonstrate that molecular crowding can exert opposite effects on the elementary equilibria involved in ion-pair extraction and clarify the different contributions of excluded volume and osmotic pressure to association and phase distribution.