Nuclear singlet states may display lifetimes that are an order of magnitude greater than conventional relaxation times. Existing methods for accessing these long-lived states require a resolved chemical shift difference between the nuclei involved. Here, we demonstrate a new method for accessing singlet states that works even when the nuclei are almost magnetically equivalent, such that the chemical shift difference is unresolved. The method involves trains of 1801 pulses that are synchronized with the spin-spin coupling between the nuclei. We demonstrate experiments on the terminal glycine resonances of the tripeptide alanylglycylglycine (AGG) in aqueous solution, showing that the nuclear singlet order of this system is long-lived even when no resonant locking field is applied. Variation of the pulse sequence parameters allows the estimation of small chemical shift differences that are normally obscured by larger J-couplings.

We review experimental techniques in our laboratory for nuclear magnetic resonance (NMR) in zero and ultralow magnetic field (below 0.1 μT) where detection is based on a low-cost, non-cryogenic, spin-exchange relaxation free Rb atomic magnetometer. The typical sensitivity is 20-30 fT/Hz for signal frequencies below 1 kHz and NMR linewidths range from Hz all the way down to tens of mHz. These features enable precision measurements of chemically informative nuclear spin-spin couplings as well as nuclear spin precession in ultralow magnetic fields.

The strong enhancement of NMR signals achieved by hyperpolarization decays, at best, with a time constant of a few minutes. Here, we show that a combination of long-lived singlet states, molecular design, magnetic field cycling, and specific radiofrequency pulse sequences allows repeated observation of the same batch of polarized nuclei over a period of 30 min and more. We report a recycling protocol in which the enhanced nuclear polarization achieved by dissolution-DNP is observed with full intensity and then returned to singlet order. MRI experiments may be run on a portion of the available spin polarization, while the remaining is preserved and made available for a later use. An analogy is drawn with a "spin bank" or "resealable container" in which highly polarized spin order may be deposited and retrieved.

Hyperpolarized singlet order is available immediately after dissolution DNP, avoiding need for additional preparation steps. We demonstrate this procedure on a sample of [1,2− 13 C 2 ]pyruvic acid.T he large signal improvement provided by spin hyperpolarization methods has dramatically extended the in vivo potential of NMR spectroscopy.1 Hyperpolarized nuclear spin order permits tracing of both endogenous and nonendogenous substances as they are transported through the blood vessels and organs and participate in metabolism.2−4 A limitation of these techniques is the often short observation time scale imposed by nuclear spin relaxation. Hyperpolarized magnetization typically decays with the longitudinal relaxation time T 1 , which is typically between a fraction of a second and 1 min and can be particularly short in vivo because of the presence of paramagnetic species and high concentrations of other magnetic nuclei. Short decay times hinder the use of hyperpolarized substrates for imaging metabolism in vivo because of the relatively long times taken for substrates to reach the tissue of interest from the point of injection. For instance, while hyperpolarized 129 Xe can be used as a lung imaging agent in gas-phase magnetic resonance imaging (MRI) because its relaxation time is several hours in the gas phase, its potential as a tracer in blood is very limited because the T 1 relaxation time of 129 Xe is reduced to only a few seconds upon dissolution. 5A potential remedy for the limited hyperpolarization lifetime is the use of nuclear singlet states involving coupled spin-1 / 2 pairs. The lifetime of the nuclear singlet state (|αβ⟩ − |βα⟩)/ √2 is capable of exceeding T 1 in some circumstances, since many common relaxation mechanisms are symmetric with respect to spin exchange and cannot induce singlet−triplet transitions. 6−9 In previous work, we reported the potential of doubly 11In some cases, substances exhibiting hyperpolarized singlet order are accessible directly by chemical reactions of hydrogen enriched with the para-hydrogen spin isomer.12−14 An alternative approach is to prepare hyperpolarized magnetization using a method such as dynamic nuclear polarization (DNP) 15,16 and then to convert it into singlet order. Several conversion methods are available:• application of resonant radiofrequency pulses in a high magnetic field to excite a "precursor state", 7 which is transformed into singlet order upon adiabatic transport to a low field; 17,18• preparation of singlet order in a high magnetic field using a radiofrequency pulse sequence;19−21• application of an audio-frequency pulse sequence in a low magnetic field on a prepolarized sample; 22• exploitation of chemical symmetry-switching reactions. 23All of these rely upon manipulations in addition to spin hyperpolarization. In certain cases they may be difficult to implement, require extra hardware, or take up valuable time during an experiment.In this communication, we show that these preparations may be circumvented by the availability of sing...