Figure 3.

Quantification of the concentrations of the paramagnetic cosolutes and measurements of their relaxivities. (A) Determination of Gd-chelate concentrations by the Evans method using coaxial NMR tubes for a high-field NMR spectrometer.³⁸ Because Gd governs the overall molar magnetic susceptibility of the Gd chelates, the Gd concentration can be determined from the experimentally observed magnetic susceptibility shift Δ in ppm using eq 1. Concentrations of the nitroxide-based cosolutes are obtained as described previously.²⁰ (B) ¹H relaxivity data for the Gd chelates and the PROXYL derivatives, focusing on longitudinal relaxation of water ¹H spins as a function of the concentration of paramagnetic cosolutes. Data were measured at a ¹H frequency of 500 MHz using a buffer of 25 mM MES (pH 5.5) and 1 mM DSS at 25 °C using a saturation-recovery method described by Toyama et al.²³ The second term of the regression was introduced to account for slight deviation from linearity. Water ¹H relaxivity data allow for calibration of the paramagnetic cosolute concentration in biomolecular samples in the same buffer.