NMR chemical shift references for binding constant determination in aqueous solutions

For deuterium oxide solutions of various compounds including cyclic and acy clic saccharides, oligoglycines, and sodium chloride., the proton chemical shifts of these solutes and the tetramethylammonium ion (TMA), referred to an external standard, change linearly with the molarity of solute. This slo pe is almost independent of the kind of protons of solute and TMA and is pr oportional to the product of the molar volume of the solute and the differe nce in volume magnetic susceptibility between deuterium oxide and the solut e. The equilibrium formation constant and structure of ion-pair of TMA and the benzenesulfonate ion (BS) are evaluated from chemical shifts referred t o an internal standard. At least some of the four methyl groups of TMA in t he ion-pair complex are located above the benzene ring of BS. This structur e indicates that TMA and BS form their ion pair by hydrophobic and electros tatic attraction. These results serve not only for choosing appropriate int ernal references and internal or external standard methods for chemical shi ft measurements in aqueous solutions but also for correcting the chemical s hift referred to the external standard.