Optimization of dynamic nuclear polarization experiments in aqueous solution at 15 MHz/9.7 GHz: a comparative study with DNP at 140 MHz/94 GHz

Abstract

Dynamic nuclear polarization is emerging as a potential tool to increase the sensitivity of NMR aiming at the detection of macromolecules in liquid solution. One possibility for such an experimental design is to perform the polarization step between electrons and nuclei at low magnetic fields and then transfer the sample to a higher field for NMR detection. In this case, an independent optimization of the polarizer and detection set ups is required. In the present paper we describe the optimization of a polarizer set up at 15 MHz (1)H NMR/9.7 GHz EPR frequencies based on commercial hardware. The sample consists of the nitroxide radical TEMPONE-D,(15)N in water, for which the dimensions were systematically decreased to fit the homogeneous B(1) region of a dielectric ENDOR resonator. With an available B(1) microwave field up to 13 G we observe a maximum DNP enhancement of -170 at room temperature by irradiating on either one of the EPR lines. The DNP enhancement was saturated at all polarizer concentrations. Pulsed ELDOR experiments revealed that the saturation level of the two hyperfine lines is such that the DNP enhancements are well consistent with the coupling factors derived from NMRD data. By raising the polarizing field and frequencies 10-fold, i.e. to 140 MHz (1)H/94 GHz EPR, we reach an enhancement of -43 at microwave field strengths (B(1) approximately 5 G). The results are discussed in view of an application for a DNP spectrometer.

Spin‐Labeled Heparins as Polarizing Agents for Dynamic Nuclear Polarization

“…The attained enhancement E = À112 for TEMPOL has been reported recently in several publications [8][9][10][11] and does not include new information, but it demonstrates the reliability of our experimental setup. Similar enhancements for SL-heparin of up to E = À110 validate the assumption that the electronnuclear coupling is mainly of dipolar origin.…”

“…[5,8] They are added to the sample as spin probes yielding the highest observed enhancements, for example, E = À170 at 0.35 T and E = À29 at 10 T at concentrations of about 20 mm. [11,12] However, for the investigation of biomolecules and proteins via DNP, a lower radical concentration is beneficial because radicals lead to NMR-line broadening and fast T 1n relaxation.…”

Saturation factor of nitroxide radicals in liquid DNP by pulsed ELDOR experiments

“…[8][9][10][11][12] Due to the strong hyperfine coupling between the electron and the nitrogen nucleus, which splits the EPR spectrum into two ( 15 N) or three ( 14 N) separate lines, the saturation factor, defined as s eff = (S B À hS z i)/S B with S B the Boltzmann polarization of the electron spin, cannot be simply extracted from the saturation behaviour of the pumped line according to the Bloch equation, as formerly suggested for trityl radicals. 13 Moreover, the expectation value of the electron spin polarization hS z i depends on the population of all energy levels involved.…”

“…the ratio of the gyromagnetic constants of the electron spin g s and the target nucleus g I , the coupling factor x, the leakage factor f and the effective saturation factor s eff of the EPR line: 6 e = 1 À s eff fx|g s |/g I (1) In recent studies, nitroxide radicals have been favoured as polarizing agents for DNP since they are soluble in water, well compatible with biological systems, non-toxic and have been found to account for large DNP enhancements at magnetic fields up to 9 T. 7,8 However, the determination of the saturation factor for this class of polarizers has emerged as one of the major difficulties in rationalizing the observed DNP enhancements in terms of the Overhauser equation (1). [8][9][10][11][12] Due to the strong hyperfine coupling between the electron and the nitrogen nucleus, which splits the EPR spectrum into two ( 15 N) or three ( 14 N) separate lines, the saturation factor, defined as s eff = (S B À hS z i)/S B with S B the Boltzmann polarization of the electron spin, cannot be simply extracted from the saturation behaviour of the pumped line according to the Bloch equation, as formerly suggested for trityl radicals. 13 Moreover, the expectation value of the electron spin polarization hS z i depends on the population of all energy levels involved.…”

“…For this measurement, the sample dimensions were optimized (0.45 mm ID, 3 mm length) as to avoid microwave heating of the sample. 12 The maximum enhancement factors are reported in Table 1. Using these values for e, the effective saturation factors s eff and the leakage factors f from ref.…”