13C Dynamic Nuclear Polarization Using a ... - ACS Publications

Jun 20, 2017 - Department of Physics, University of Texas at Dallas, 800 West Campbell Road, .... at DNP conditions of 3.35 T and 1.2 K. W-Band EPR wa...
0 downloads 0 Views 1MB Size
Download PDF
Article pubs.acs.org/JPCA

13

C Dynamic Nuclear Polarization Using a Trimeric Gd3+ Complex as an Additive

Peter Niedbalski,† Christopher Parish,† Qing Wang,† Andhika Kiswandhi,† Zahra Hayati,‡ Likai Song,‡ and Lloyd Lumata*,† †

Department of Physics, University of Texas at Dallas, 800 West Campbell Road, Richardson, Texas 75080 United States National High Magnetic Field Laboratory, Florida State University, 1800 E. Paul Dirac Drive, Tallahassee, Florida 32310, United States

‡

S Supporting Information *

ABSTRACT: Dissolution dynamic nuclear polarization (DNP) is one of the most successful techniques that resolves the insensitivity problem in liquid-state nuclear magnetic resonance (NMR) spectroscopy and imaging (MRI) by amplifying the signal by several thousandfold. One way to further improve the DNP signal is the inclusion of trace amounts of lanthanides in DNP samples doped with trityl OX063 free radical as the polarizing agent. In practice, stable monomeric gadolinium complexes such as Gd-DOTA or Gd-HPDO3A are used as beneficial additives in DNP samples, further boosting the DNP-enhanced solid-state 13C polarization by a factor of 2 or 3. Herein, we report on the use of a trimeric gadolinium complex as a dopant in 13C DNP samples to improve the 13C DNP signals in the solid-state at 3.35 T and 1.2 K and consequently, in the liquid-state at 9.4 T and 298 K after dissolution. Our results have shown that doping the 13C DNP sample with a complex which holds three Gd3+ ions led to an improvement of DNP-enhanced 13C polarization by a factor of 3.4 in the solid-state, on par with those achieved using monomeric Gd3+ complexes but only requires about one-fifth of the concentration. Upon dissolution, liquid-state 13 C NMR signal enhancements close to 20 000-fold, approximately 3-fold the enhancement of the control samples, were recorded in the nearby 9.4 T high resolution NMR magnet at room temperature. Comparable reduction of 13C spin−lattice T1 relaxation time was observed in the liquid-state after dissolution for both the monomeric and trimeric Gd3+ complexes. Moreover, W-band electron paramagnetic resonance (EPR) data have revealed that 3-Gd doping significantly reduces the electron T1 of the trityl OX063 free radical, but produces negligible changes in the EPR spectrum, reminiscent of the results with monomeric Gd3+complex doping. Our data suggest that the trimeric Gd3+ complex is a highly beneficial additive in 13C DNP samples and that its effect on DNP efficiency can be described in the context of the thermal mixing mechanism.

■

INTRODUCTION Dynamic nuclear polarization (DNP) via the dissolution process is one of the most recent and successful methods used to solve the problem of low signal sensitivity in nuclear magnetic resonance (NMR) spectroscopy and imaging (MRI). This NMR signal amplification technique works by transferring the high spin alignment from electrons, which have much higher polarization than the nuclei under identical conditions, to nuclei by microwave irradiation near the electron paramagnetic resonance (EPR) frequency.1,2 This polarization transfer process is most effective at cryogenic temperatures where the electron Boltzmann thermal polarization approaches 100%.3 Samples polarized at such conditions may then be converted to highly polarized liquid-state samples by way of dissolution technique.4,5 In this process, after hyperpolarization in the solid state, superheated water or other solvent is used to rapidly dissolve the sample, maintaining the high level of polarization as the sample is transferred as a liquid from the polarizer to a high-resolution NMR magnet or MRI scanner for © XXXX American Chemical Society

detection. As a consequence, detection or imaging of nonproton or low-gyromagnetic ratio (γ) nuclei at mM concentrations has been made possible with this technology with excellent sensitivity.4,6−9 Specifically, the dissolution DNP technology has been applied to numerous biomedical applications including in vitro and in vivo 13C metabolic imaging and spectroscopy.10−25 The efficiency of DNP depends upon a number of factors, one of the most important being the choice of free radical that supplies the free electrons from which polarization is transferred.26−31 For instance, numerous experiments4,32−37 have shown that the narrow-EPR line width free radical trityl OX063 (see structure in Figure 1) is an especially effective polarizing agents for low-γ nuclei such as 13C spins. On the other hand, a number of comparative reports34−37 have shown Received: April 25, 2017 Revised: June 19, 2017 Published: June 20, 2017 A

DOI: 10.1021/acs.jpca.7b03869 J. Phys. Chem. A XXXX, XXX, XXX−XXX

Article

The Journal of Physical Chemistry A

Figure 1. Gadolinium complexes and the free radical polarizing agent used in this study.

DNP. The influence of 3-Gd doping on 13C DNP was optimized and tested on our model compound [1-13C] acetate at DNP conditions of 3.35 T and 1.2 K. W-Band EPR was performed to shed light on the effect of 3-Gd doping on relaxation properties of trityl OX063 electrons. Furthermore, liquid-state NMR signal enhancements were quantified and compared with the DNP results obtained with monomeric GdDOTA and reference samples.

that nitroxide-based free radicals such as TEMPO can also hyperpolarize 13C spins though to a lesser degree than trityl OX063, owing to their significantly larger EPR line widths which are better matched for polarizing large-γ nuclei such as 1 H spins.38−40 Aside from the choice of free radicals, the location of 13C isotopic labeling in a compound of interest can also affect the maximum achievable 13C DNP levels.41 On another note, isotopic enrichment (e.g., 2H and 13C) of the glassing matrix39,42,43 can, under certain conditions, also have beneficial effects on 13C DNP in terms of higher 13C polarization level and faster polarization buildup. These examples suggest that modification of the DNP sample components can have significant impact on the efficiency of DNP. Additionally, doping the DNP sample with trace amounts of lanthanide ions can enhance the 13C DNP signals further.34,44−47 This has been shown most extensively using gadolinium complexes due both to their wide availability as contrast agents in MRI and to the success of gadolinium complexes as beneficial additives in DNP.34,35,44 In addition, other lanthanides including holmium, terbium, and dysprosium have also been shown to be effective for improving 13C polarization levels achieved in DNP.46,47 It should be noted that these lanthanide additives are not the DNP polarizing agents themselves, but the improvement in 13C DNP signals emanates from the interaction of these lanthanides with the electronic properties of the stable organic free radical used in the experiments. These lanthanides ions are most often chelated within a stable ligand in order to render them inert if used in vivo in biomedical applications. One such ligand that has seen considerable use is DOTA, which shows considerable thermal and chemical stability.48,49 So far, each of the ligands commonly used for lanthanide-enhanced DNP holds only a single Ln(III) ion. In this work, we have explored the use of an additive in DNP sample in which three DOTA ligands are tethered together to hold three Gd3+ ions within a single complex (see Figure 1). This trimeric gadolinium complex (hereafter known as 3-Gd) has been used as a DNP enhancer in several in vitro and in vivo dissolution DNP studies;17−25 however, there have been no extensive studies as to its detailed optimization and physical characterization. Thus, the main goal of this study was to evaluate and elucidate the effect of 3-Gd doping on 13C

■

EXPERIMENTAL SECTION Sample Preparation. The trityl OX063 free radical (Oxford Instruments, MA), [1-13C] sodium acetate (Cambridge Isotope Laboratories, MA), Gd-DOTA (Macrocyclics Inc., Plano, TX), and glassing solvents (Sigma-Aldrich, MO) used in this study were obtained commercially and were used without further purification. 3-Gd (MW = 2108.47 g/mol) was received as a gift from Albeda Research (Copenhagen, Denmark) and was also used without further purification. Each DNP sample was made with 24.9 mg of [1-13C] sodium acetate and 2.14 mg trityl OX063 dissolved in a glassing solution of 100 μL 1:1 v/v water:glycerol doped with 3-Gd. The final concentrations of [1-13C] sodium acetate and trityl OX063 free radical in each DNP sample were 3 M and 15 mM, respectively. The glassing solution was prepared by creating a 2 mM stock solution of 3-Gd in 1:1 v/v water/glycerol and diluting to the appropriate concentration using a stock solution of pure 1:1 v/v water/glycerol. In addition, the same sample formulation was used to prepare 100 μL aliquots of 2 mM GdDOTA for comparison with the 3-Gd DNP samples. All DNP samples were made a few hours before the 13C DNP experiments and were stored in −80 °C refrigerator (ThermoFisher Scientific, WI) prior to the DNP experiments. 13 C Microwave Frequency Sweeps. All DNP experiments were performed at the Advanced Imaging Research Center (AIRC) at the University of Texas Southwestern (UTSW) Medical Center on a HyperSense polarizer (Oxford Instruments, UK). This instrument operates using a magnetic field of 3.35 T and a sample space with a base temperature of 1.2 K when the roots blower pump (Edwards Vacuum, UK) is turned on. Aliquots (100 μL) of 3 M [1-13C] sodium acetate in 1:1 v/v glycerol:water doped with 15 mM trityl OX063 and with different concentrations of 3-Gd were inserted in the B

DOI: 10.1021/acs.jpca.7b03869 J. Phys. Chem. A XXXX, XXX, XXX−XXX

Article

The Journal of Physical Chemistry A polarizer for 13C microwave frequency sweeps. Sweeps were measured using steps of 5 MHz in the range 94.05−94.20 GHz with a 90 s duration time of microwave irradiation for each data point. This microwave frequency tuning range covers the full profile of 13C DNP spectra of the samples doped with trityl OX063. The locations of the optimum microwaves irradiation frequencies, namely, the positive P(+) and negative P(−) polarization peaks, were determined for each sample. Sweep data with 1 h microwave irradiation time near the P(+) of the 3-Gd doped sample are shown in Figure S1 in the Supporting Information. Solid-State 13C DNP Buildup Curves. After the optimum microwave frequencies were located from the microwave sweep data, the 13C DNP samples were irradiated at their respective optimum microwave frequencies that correspond to P(+). Various concentrations of 3-Gd doping ranging from 0 to 2 mM in 100 μL aliquots of DNP samples were tested by recording their 13C DNP signal buildup curves in the HyperSense polarizer at 3.35 T and 1.2 K. The polarization buildup monitoring was set up using the HyperSense control software wherein a shallow flip-angle RF pulse close to 2 degrees was applied every 3 min until the curves reach their maximum values.41 The least concentration of 3-Gd that provided the highest polarization level was selected as the optimum concentration for 13C DNP. The 13C polarization buildup curves for the control sample (0 mM 3-Gd) and samples doped with optimum concentration of 3-Gd and GdDOTA were done in triplicate. Mean values and standard deviations of the relative solid-state 13C polarization levels were calculated and plotted as bar graphs. Liquid-State 13C NMR Enhancements and T1 Decays. After reaching the maximum 13C polarization levels at 3.35 T and 1.2 K, the aforementioned 13C DNP samples (control, 0.4 mM 3-Gd, 2 mM Gd-DOTA) were quickly dissolved with superheated water and shuttled into a 9.4 T high-resolution NMR magnet at 298 K using a standard procedure for dissolution in the HyperSense polarizer. The dissolution medium used was 4 mL deionized water and the shuttling time of the dissolution liquid from the polarizer to the NMR magnet was approximately 8 s.8,41 A 10 mm NMR tube (Wilmad-LabGlass, NJ) already placed in a 400 MHz highresolution Varian NMR magnet (Agilent Technologies, CA) was used to collect the hyperpolarized liquid for NMR detection. To monitor liquid-state 13C T1 decay, a 13C NMR spectrum was recorded every 2 s using a 2-degree RF pulse. This measurement was initiated immediately after the completion of the dissolution liquid transfer to the NMR tube. The 13C NMR signal decay data in the liquid-state were then fitted with an equation that includes the effect of both RF pulsing and 13C T1 relaxation.8,41,50 The liquid-state 13C T1 values were calculated and plotted in the graph. To calculate the 13C NMR signal enhancement, a thermal 13C NMR signal of the liquid sample was recorded by applying a 90° RF pulse with 4 scans. The ratio of the integrated 13C NMR areas of the first hyperpolarized 13C spectrum on the T1 decay data over the thermal 13C NMR signal was calculated. The 13C NMR signal enhancement in the liquid-state was quantified using this ratio and the corrections accounting for the number of scans and the different RF flip angles used. Following the pattern of the relative solid-state 13C DNP results, average values and standard deviations (N = 3 trials) of the liquid-state 13C NMR signal enhancement numbers of the three 13C DNP samples of

interest (control, 0.4 3-Gd, and 2 mM Gd-DOTA) were calculated and plotted for comparison. W-Band EPR Measurements. W-Band (95 GHz) EPR was performed on the three DNP samples of interest (0 mM 3-Gd, 0.4 mM 3-Gd, and 2 mM Gd-DOTA) each containing 3 M [1-13C] sodium acetate and 15 mM trityl OX063 in 1:1 glycerol:water. These experiments were performed at the National High Magnetic Field Laboratory (NHMFL) in Tallahassee, FL on a Bruker E680 EPR spectrometer (Bruker Biospin, Billerica, MA) using a Bruker TE011 cylindrical cavity. Before inserting into the cavity, samples were loaded in 0.15 mm ID thin quartz capillary tubes. The sample temperature was regulated within a CF1200 helium flow cryostat (Oxford Instruments, UK). The temperature dependence of trityl OX063 electron T1 was measured between 5 and 200 K using saturation recovery method. EPR spectra were recorded using the field-swept electron spin−echo method.30 EPR spectra of these samples at 7.5 K were plotted for comparison in addition to the graph of trityl OX063 electron relaxation rate T1−1 vs temperature data. Data Analyses. Liquid-state NMR data were analyzed using VNMRJ software (Agilent Technologies, CA) and ACD/ Laboratories version 12.0 (Advanced Chemistry Development). All other data was analyzed using Igor Pro version 6.37 (Wavemetrics, Lake Oswego, OR). The modified Borghini modeling of the 13C DNP spectra (Figure S1 in the Supporting Information) was done using MatLab (Mathworks, Nattick, MA).

■

RESULTS AND DISCUSSION Several studies7,8,51,52 have indicated that, at least in the DNP conditions of 3.35 T and temperatures close to 1 K, 13C samples doped with trityl OX063 free radical are polarized predominantly via the thermal mixing DNP mechanism. In thermal mixing or equal spin temperature (EST) theory, a thermal contact is established between the electron dipolar system (EDS) and nuclear Zeeman system (NZS) due to their comparable energies.1−3,31,53,54 Microwave irradiation at a frequency that is slightly off-resonance dynamically cools the electrons which in turn cools the nuclear spins, eventually reaching a common spin temperature that is lower than the lattice temperature.1−3,31,53,54 In other words, the high spin alignment or polarization of the electrons is being mimicked by the nuclear spins translating to highly enhanced NMR signals relative to the signal acquired at thermal equilibrium. Other possible mechanisms that may come into play are the solid effect (SE)55 and the cross effect (CE).56−59 The SE is a twospin mechanism in which nuclear and electron spins are coupled and polarization proceeds by way of microwave excitation of forbidden transitions.55 This mechanism dominates when the free radical EPR line width is narrower than the nuclear Larmor frequency. The CE is a three-spin mechanism that is only effective when two electron spins S1 and S2 and a nuclear spin I are related such that ωI = ωS1 − ωS2.56−59 In practice, this may take place when the EPR line width of the free radical is inhomogeneously broadened to greater than the nuclear Larmor frequency. When the cross effect condition is met, a degeneracy is introduced causing irradiation at the frequency of one of the electrons to excite the forbidden transition of the other electron and the nucleus. We will revisit this discussion of possible DNP mechanisms later with the DNP and EPR results. C

DOI: 10.1021/acs.jpca.7b03869 J. Phys. Chem. A XXXX, XXX, XXX−XXX

Article

The Journal of Physical Chemistry A In order to effectively track and optimize the effect of 3-Gd doping on 13C DNP, we have first performed 13C microwave DNP sweeps of 13C acetate samples doped with various 3-Gd concentrations at 3.35 T and 1.2 K. This is an important step because the optimum locations of the microwave irradiation frequencies, namely, the positive P(+) and negative P(−) polarization peaks, could significantly change with Gd3+ doping as was observed with Gd-DOTA and other monomeric lanthanide complexes.8,34,35,44−47 Due to the large number of samples for optimization studies, the sweep data here were taken with a relatively short irradiation time (90 s) for each point in the DNP spectrum. Since polarization buildup rate constants and asymptotic values are dependent upon the microwave irradiation frequencies, a concern might arise whether such a fast method would yield the actual DNP spectra were it done with a longer irradiation time or full polarization buildup for each frequency point. This issue was previously resolved in a previous published extensive study34 in which the main finding was while there were subtle or slight differences in the shape of DNP spectra between the fast microwave freq sweep (e.g., 3 min irradiation time) and the longer irradiation time sweep (e.g., 1 h), the locations of the P(+) and P(−) were essentially the same. To make a quick verification, additional measurements (Figure S1, Supporting Information) of DNP spectrum with longer polarization buildup times of 1 h were performed near the P(+) peak of the optimally doped (0.4 mM 3-Gd) sample. These data reveal that the locations of the P(+) obtained with a 90-s and 1-h irradiation time step measurement are practically the same, corroborating with the results of the previously published 13C DNP spectra data between the control and Gd3+-doped samples.34 Inspection of Figure 2 reveals that even the smallest concentration of 3-Gd studied led to significant narrowing of the DNP spectrum similar to that seen when using monomeric lanthanide complexes.34,35,46,47 For the control sample without any lanthanide additives, the P(+) and P(−) polarization peaks are separated by about 105 MHz. With the addition of even 0.1 mM 3-Gd, this separation is reduced to about 70 MHz. At the optimum concentration of 3-Gd (0.4 mM), the peaks are 55 MHz apart and come to a minimum separation of 40 MHz with 2 mM 3-Gd added. In comparison, the 13C microwave DNP spectrum of optimally doped Gd-DOTA DNP sample is similar to that of 3-Gd at 0.4 mM, with a separation distance between P(+) and P(−) of 60 MHz. Once the optimum microwave irradiation frequencies were determined, relative solid-state 13C polarization buildup curves of 100 μL aliquots of 3 M [1-13C] acetate samples with various 3-Gd concentrations were monitored and recorded at 3.35 T and 1.2 K as shown in Figure 3a. For consistency, all 13C DNP samples were polarized at their corresponding P(+) irradiation frequencies. Normalized with respect to the DNP signal of the control sample, a plot of the maximum solid-state 13C DNP signals of samples doped with various 3-Gd concentrations is shown in Figure 3b. It is apparent in Figure 3b that the optimum 3-Gd concentration for 13C DNP is approximately 0.4 mM. This optimum concentration is significantly less, although expected since the 3-Gd molecule holds three Gd-DOTA complexes, compared to the optimum concentration of GdDOTA at 2 mM. As shown in Figure 3a and b, samples doped with any amount of 3-Gd up to 2 mM gained solid-state 13C polarization enhancements significantly greater than that achieved by the reference sample. At 0.1 mM, the polarization enhancement was already about 2.5 times that of the reference

Figure 2. Stacked plot of microwave frequency sweeps of relative 13C DNP signals of [1-13C] sodium acetate doped with 15 mM trityl OX063 and with varying concentrations of 3-Gd. The long dashed arrow points to the direction of increasing 3-Gd concentration. The short up and down arrows indicate the positive P(+) and negative P(−) polarization peaks, respectively. These data were taken at 3.35 T and 1.2 K.

sample and at the optimum concentration of 0.4 mM, it improved to a factor of 3.4-fold relative to the 13C DNP signal of the control sample. From there, it decreased slightly to about 3.25-fold increase until a concentration of 0.8 mM, then more significantly down to about 2.75-fold. Figure 3c displays a summary of the maximum solid-state 13C DNP signals achieved with the control sample and samples doped with 0.4 mM 3-Gd and 2 mM Gd-DOTA. Both 3-Gd and Gd-DOTA doping have significantly improved 13C DNP by a factor close to 3.5-fold relative to the control sample DNP signal. Overall, it is apparent that the efficiency of 3-Gd as a beneficial additive in 13 C DNP samples is on par with that of the monomeric Gd3+ complex Gd-DOTA but it only requires about one-fifth of the concentration compared to the latter. So far, 3-Gd doping in 13C samples has shown significant effects in DNP, namely the narrowing of the 13C microwave DNP spectra and the substantial improvement in the 13C DNP signalseffects that are reminiscent of the DNP behavior with monomeric Gd3+ complex doping.34,35,46,47 To shed light on these physical behavior, we have performed W-band (3.35 T/ 95 GHz) EPR spectral and relaxation measurements on the 13C DNP samples as shown in Figure 4. Figure 4a displays the representative field-swept W-band EPR spectra of 15 mM trityl OX063 of the control, 3-Gd (0.4 mM), and Gd-DOTA (2 mM) DNP samples taken at 7.5 K. The EPR spectral measurement was focused on trityl OX063 resonance because the observed DNP behavior is ascribed to the effect of lanthanide doping on the physical properties of the free radical polarizing agent. Inspection of Figure 4a reveals that 3-Gd doping, similar to Gd-DOTA doping, at the optimum D

DOI: 10.1021/acs.jpca.7b03869 J. Phys. Chem. A XXXX, XXX, XXX−XXX

Article

The Journal of Physical Chemistry A

Figure 4. W-Band EPR results of 13 C DNP samples: (a) Representative field-swept EPR spectra of trityl OX063 for the control sample and samples doped with 3-Gd (0.4 mM) and Gd-DOTA (2 mM) taken at 7.5 K. The black curves on the EPR spectra are fits to the Voigt equation. (b) Log−log plot of the temperature dependence of trityl OX063 electron relaxation rate T1−1 for control, 3-Gd, and Gd-DOTA DNP samples. The dashed lines are fits to a power-law equation in which α is the exponent.

Figure 3. Solid-state 13C DNP results of frozen samples of 3 M [1-13C] acetate in 1:1 v/v glycerol/water with 15 mM trityl OX063: (a) Representative 13C polarization buildup curves as a function of microwave irradiation time for 13C acetate samples with varying concentrations of 3-Gd. (b) Plot of the relative maximum 13C DNP signals achieved with various 3-Gd concentrations. These data points were normalized with the 13C DNP signal of the control sample (0 mM 3-Gd). The up arrow indicates the approximate optimum concentration of 3-Gd (∼0.4 mM) for 13C DNP. (c) Summary of the relative maximum 13C DNP signals of the control sample and samples doped with optimum concentrations of 3-Gd (0.4 mM) and GdDOTA (2 mM). These results are given as average values and the errors bars are standard deviations with N = 3 trials.

component is ascribed to electron−electron cross relaxation effects and the longer relaxation component corresponds to the actual electron T1 relaxation.59 The temperature-dependent trityl OX063 relaxation rate data in Figure 4b were fitted to a power law equation T1−1 = ATα, where A is a constant and α is the exponent that could suggest the relaxation mechanism of the electrons.52 Above 80 K, the relaxation rates for all three samples (control, 3-Gd, Gd-DOTA) were nearly identical and overlapping with values of α suggesting a combination of Raman and other multiphonon electron relaxation process.52 Below 80 K, the trityl OX063 electron relaxation rates of samples doped with 3-Gd and Gd-DOTA start to separate from the T1−1 values of the control sample and this behavior becomes more prominent as the temperature drops close to the base temperature of 5 K. At low temperatures below 20 K, the slopes of relaxation rate values of these three samples in the log−log scale are close to 1, suggesting the one-phonon direct process60 as the predominant electron relaxation mechanism in this temperature regime for the control and lanthanide-doped samples. Between 5 and 65 K, the trityl OX063 electron relaxation rates of samples doped with 0.4 mM 3-Gd almost overlap with the relaxation rate data of DNP samples doped with 2 mM Gd-DOTA. At the base temperature of 5 K, both the monomeric and trimeric lanthanide-doped samples have relaxation rates that are approximately an order of magnitude higher than that of the control sample.

concentration has a negligible effect on the EPR spectrum of trityl OX063. The corresponding full width at half-maximum (fwhm) values of trityl OX063 EPR spectra displayed in Figure 4a for the control and samples doped with Gd-DOTA and 3Gd are 48, 47, and 45 MHz, respectively. These EPR results imply that the narrowing of the 13C microwave DNP spectra with 3-Gd doping cannot be adequately explained by the almost exclusive reliance of the Borghini model54 on the shape of the EPR spectrum since the latter was essentially unchanged with lanthanide doping. While the EPR spectrum of trityl OX063 was not significantly affected by 3-Gd doping, its electron T1 relaxation was significantly changed with this additive. Figure 4b displays the temperature dependence of trityl OX063 electron relaxation rate T1−1 at W-band for the control 13C DNP sample and 13C DNP samples doped with the optimum concentrations of 3-Gd (0.4 mM) and Gd-DOTA (2 mM). EPR relaxation measurements were performed as described previously.46,47 This included fitting the electron relaxation recovery curves with a double-exponential function in which the shorter relaxation E

DOI: 10.1021/acs.jpca.7b03869 J. Phys. Chem. A XXXX, XXX, XXX−XXX

Article

The Journal of Physical Chemistry A

potentially f are changed with the addition of 3-Gd or GdDOTA. Previous research has shown that the 13C nuclear relaxation time of [1-13C] pyruvate is essentially unchanged or minimally reduced with doping of optimum concentrations of monomeric lanthanide complexes,34 which suggests that the same will be true for [1-13C] acetate given the similar molecular environments present. Using this assumption, η in eq 1 is drastically reduced by the shortening of trityl OX063 electron T1 with 3-Gd or Gd-DOTA doping, hence leading to an increase in polarization. Thermodynamically speaking, the reduced η translates to faster cooling rate of the electron Zeeman system than the heating rate of the electron dipolar system via the interaction of the latter with the lattice.31 Since the microwave irradiation establishes a thermal contact between the two systems, the electron dipolar system is then cooled by the electron Zeeman system.31 An electron dipolar system with a lower spin temperature is quite beneficial for the nuclear Zeeman system as it acquires the same spin temperature via the thermal mixing process, leading to higher nuclear polarization or enhanced NMR signals. Thus, the improvement of 13C DNP signals with 3-Gd doping is attributed to the reduced electron T1 as well in the context of thermal mixing DNP mechanism. The enhanced solid-state 13C polarization levels acquired at cryogenic temperatures were mostly preserved in the liquidstate after dissolution with water as revealed in Figure 5. The liquid-state 13C NMR signal enhancements reported here were calculated from hyperpolarized 13C NMR spectra recorded with a 2° RF pulse immediately after the dissolution liquid transfer. The transit of the dissolution liquid from the hyperpolarizer to the high-resolution NMR magnet takes about 8 s to complete.8,46,47 Representative hyperpolarized and thermal 13 C NMR spectra are displayed in Figure 5a. The NMR signal enhancements were calculated using an equation that accounts for the ratio of integrated areas of the hyperpolarized and thermal NMR signals and corrections due to differences in the RF pulse angles used.8,50 Despite the losses due to T1 decay and the journey through near zero magnetic field areas during the dissolution liquid transit,68 liquid-state 13C NMR signal enhancements close to 20 000-fold were recorded for the 3Gd doped samples at 9.4 T and 298 K as shown in Figure 5b. The liquid-state 13C NMR signal enhancements for both 3-Gd and Gd-DOTA shown in Figure 5b were approximately three times that of the reference sample. These liquid-state NMR enhancement results are in close proportion to the relative solid-state 13C DNP levels of these samples obtained at cryogenic temperature. Despite a fewer number of Gd3+ ions required for efficient 13C DNP using the more compact 3-Gd complex, the 13C T1 relaxation decay of samples doped with 3Gd was found to be similar to that of Gd-DOTA sample dissolution liquid as shown in Figure 5c. This is somewhat surprising considering that 3-Gd dissolution liquid contains only 3/5 of the total Gd3+ ions compared to the Gd-DOTA dissolution liquid. However, it is likely that the large 3-Gd molecule has comparatively slow molecular tumbling time, increasing the relaxivity of the contained Gd3+ ions thus increasing the 13C relaxation rate.69−71 Nevertheless, the overall results shown here suggest that the more compact 3-Gd is a highly beneficial additive in 13C DNP samples that yields comparable 13C DNP improvements observed with the monomeric Gd3+ complexes but with less optimum concentration required.

The reduction in the trityl OX063 electron T1 is by far the most striking effect of 3-Gd doping observed in the EPR measurements. We attribute the observed DNP effects of narrowing of 13C microwave DNP spectra and increased 13C polarization to the drastic decrease in electron T1 relaxation of trityl OX063 with 3-Gd doping. First, we would like to comment on the narrowing of the 13C DNP spectra with 3-Gd doping. It can be seen from Figure 2 that the separation distance between P(+) and P(−) was decreased by about 45 MHz with the addition of 0.4 mM 3-Gd. However, as mentioned before, there was no visible change or narrowing in the trityl OX063 EPR spectrum with 3-Gd doping. Thus, it must be the reduced electron T1 of trityl OX064 that must account for the narrowing of the 13C DNP spectrum. Since the original Borghini model is known to be insufficient to accurately predict the DNP spectra,61 we invoked an extended model of thermal mixing put forth by Wenckebach 62 (Supporting Information) in which both the EPR spectrum and electron T1 are taken into account in addition to other parameters to fit the DNP spectra of the control and 3-Gddoped samples. Numerical simulations using this modified Borghini model62 with the actual EPR line width and electron T1 data yielded good fits with the experimental DNP spectra for both the control and 3-Gd-doped samples, accurately predicting the P(+) and P(−) locations (Figure S1, Supporting Information) in particular. The simulation results using this thermal mixing model indicate that the narrowing of DNP spectra can be ascribed to the reduction of electron T1. This finding is in line with the other extended models of thermal mixing which suggest electron T1 as a dominant factor that can cause the observed narrowing in DNP spectra as indicated by Serra anc co-workers.63−65 Alternatively, the DNP spectra could also be potentially explained in terms of a combination of the solid effect and cross effects.66,67 As the concentration of 3Gd is increased, the contribution from the CE increases to a greater extent than that from the SE, narrowing the DNP spectrum.66,67 However, this explanation currently does not accurately describe the initial difference between the positive and negative polarization peaks. For a case of pure solid effect, the separation would be expected to be 2νI, in this case about 70 MHz.55 The addition of a cross effect component would only serve to narrow this peak separation further.56 The initial distance between P(+) and P(−) is 105 MHz and only after the addition of 3-Gd does the peak separation reach a value that could indicate solid or cross effects. Our data here, however, favor thermal mixing mechanism via reduced electron T1 to explain the narrowing of the 13C microwave DNP spectra with 3-Gd doping. Using the assumption that thermal mixing dominates the DNP process, the explanation for increased 13C polarization with the addition of 3-Gd is attributed to the reduction of the trityl OX063 electron T1 relaxation time. The theoretical maximum polarization under the thermal mixing model is given by31 ⎛ ωω Pmax = tanh⎜⎜βL e I ⎝ 4D

⎞ ⎟ ⎟ n(1 + f ) ⎠ 1

(1)

In this equation, βL is the inverse lattice temperature, ωe and ωI are the electron and nuclear Larmor frequencies, D is the free radical EPR line width, η is the ratio of nuclear to electron T1 relaxation times, and f is a “leakage factor” containing all effects from nuclear relaxation.31 Of these parameters, only η and F

DOI: 10.1021/acs.jpca.7b03869 J. Phys. Chem. A XXXX, XXX, XXX−XXX

Article

The Journal of Physical Chemistry A

DOTA, and thus could be a viable alternative as an additive in DNP sample preparation.

■

ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jpca.7b03869. Comparison of actual experimental data and numerical modeling of 13C DNP spectra (PDF)

■

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. ORCID

Lloyd Lumata: 0000-0002-3647-3753 Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS The authors would like to acknowledge support from the U.S. Department of Defense (DOD) Grant Number W81XWH-141-0048 (L.L.), as well as the Robert A. Welch Foundation Grant Number AT-1877 (L.L.). Additionally, the authors acknowledge the NHMFL user collaboration grants program Award No. 5080 (L.S.). EPR was performed at NHMFL, which is supported by the National Science Foundation (NSF) Cooperative Agreement No. DMR 1157490 and the State of Florida. DNP experiments were performed at the Advanced Imaging Research Center (AIRC) at the University of Texas Southwestern Medical Center. The DNP facility in the AIRC is supported by the National Institutes of Health (NIH) Grant Number 8P41-EB015908. The authors would also like to thank Dr. Mathilde Lerche and Dr. Magnus Karlsson of Albeda Research and Denmark Technological University for providing the 3-Gd compound used in this study.

Figure 5. Liquid-state 13C NMR results at 9.4 T and 298 K: (a) Representative hyperpolarized (2° RF pulse, 1 scan) and thermal (90° RF pulse, 4 scans) 13C NMR spectra of 73 mM aqueous solutions of 13 C acetate. (b) Plot of the average liquid-state 13C NMR signal enhancements of the control, 3-Gd (0.4 mM), and Gd-DOTA (2 mM) DNP samples calculated after dissolution with 4 mL water. The error bars are standard deviations with N = 3 trials. (c) Decay of the hyperpolarized 13C NMR signals of the aforementioned DNP samples (control, 3-Gd, and Gd-DOTA). These data were plotted by applying a 2-degree RF pulse every 2 s.

■

REFERENCES

(1) Abragam, A.; Goldman, M. Principles of Dynamic Nuclear Polarisation. Rep. Prog. Phys. 1978, 41, 395. (2) Günther, U. L. Dynamic Nuclear Hyperpolarization in Liquids. In Modern NMR Methodology; Heise, H., Matthews, S., Eds.; Topics in Current Chemistry; Springer: Berlin, Heidelberg, 2011; pp 23−69. (3) Crabb, D. G.; Meyer, W. Solid Polarized Targets for Nuclear and Particle Physics Experiments. Annu. Rev. Nucl. Part. Sci. 1997, 47, 67− 109. (4) Ardenkjær-Larsen, J. H.; Fridlund, B.; Gram, A.; Hansson, G.; Hansson, L.; Lerche, M. H.; Servin, R.; Thaning, M.; Golman, K. Increase in Signal-to-Noise Ratio of > 10,000 Times in Liquid-State NMR. Proc. Natl. Acad. Sci. U. S. A. 2003, 100, 10158−10163. (5) Ardenkjaer-Larsen, J. H. On the Present and Future of Dissolution-DNP. J. Magn. Reson. 2016, 264, 3−12. (6) Jiang, W.; Lumata, L.; Chen, W.; Zhang, S.; Kovacs, Z.; Sherry, A. D.; Khemtong, C. Sci. Rep. 2015, 5, 9104. (7) Lumata, L.; Merritt, M. E.; Malloy, C.; Sherry, A. D.; Kovacs, Z. Fast Dissolution Dynamic Nuclear Polarization NMR of 13C-enriched 89 Y-DOTA Complex: Experimental and Theoretical Considerations. Appl. Magn. Reson. 2012, 43, 69−79. (8) Lumata, L.; Jindal, A. K.; Merritt, M. E.; Malloy, C. R.; Sherry, A. D.; Kovacs, Z. DNP by Thermal Mixing under Optimized Conditions Yields > 60000-Fold Enhancement of 89Y NMR Signal. J. Am. Chem. Soc. 2011, 133, 8673−8680. (9) Lumata, L.; Merritt, M. E.; Hashami, Z.; Ratnakar, S. J.; Kovacs, Z. Production and NMR Characterization of Hyperpolarized 107,109Ag Complexes. Angew. Chem., Int. Ed. 2012, 51, 525−527.

■

CONCLUSION In conclusion, we have found that the trimeric gadolinium complex 3-Gd is a efficient and beneficial additive for increasing 13 C DNP of samples doped with the trityl OX063 free radical. Its DNP performance in both solid and liquid states is almost identical to the commonly used Gd-DOTA. Furthermore, with an optimum concentration of 0.4 mM, a smaller total number of Gd3+ ions are needed for effective polarization enhancement. It was shown that 3-Gd doping could further boost the solidstate 13C DNP signal of reference samples by a factor of 3.4. The increase in the 13C polarization levels with 3-Gd doping is ascribed to the reduction of the trityl OX063 electron T1 which results in lower spin temperature of the 13C spins. Moreover, the narrowing of the 13C DNP spectrum with 3-Gd doping can also be attributed to the reduced electron T1 via a prediction of the thermal mixing model. The overall results have shown that the more compact 3-Gd is an equally efficient DNP enhancer compared with the monomeric DNP additive such as GdG

DOI: 10.1021/acs.jpca.7b03869 J. Phys. Chem. A XXXX, XXX, XXX−XXX

Article

The Journal of Physical Chemistry A (10) Gallagher, F. A.; Kettunen, M. I.; Brindle, K. M. Biomedical Applications of Hyperpolarized 13C Magnetic Resonance Imaging. Prog. Nucl. Magn. Reson. Spectrosc. 2009, 55, 285−295. (11) Brindle, K. M.; Bohndiek, S. E.; Gallagher, F. A.; Kettunen, M. I. Tumor Imaging Using Hyperpolarized 13C Magnetic Resonance Spectroscopy. Magn. Reson. Med. 2011, 66, 505−519. (12) Day, S. E.; Kettunen, M. I.; Gallagher, F. A.; Hu, D.-E.; Lerche, M.; Wolber, J.; Golman, K.; Ardenkjaer-Larsen, J. H.; Brindle, K. M. Detecting Tumor Response to Treatment Using Hyperpolarized 13C Magnetic Resonance Imaging and Spectroscopy. Nat. Med. 2007, 13, 1382−1387. (13) Golman, K.; Petersson, J. S. Metabolic Imaging and Other Applications of Hyperpolarized 13C. Acad. Radiol. 2006, 13, 932−942. (14) Yang, C.; Ko, B.; Hensley, C.; Jiang, L.; Wasti, A.; Lumata, L.; Mitsche, M.; Merritt, M.; DeBerardinis, R. J.; Kim, J.; Sudderth, J.; Calvaruso, M. A.; Rutter, J. Glutamine Oxidation Maintains the TCA cycle and Cell Survival During Impaired Mitochondrial Pyruvate Transport. Mol. Cell 2014, 56, 414−424. (15) Lumata, L.; Yang, C.; Ragavan, M.; Carpenter, N.; DeBerardinis, R. J.; Merritt, M. E. Hyperpolarized 13C magnetic resonance and its use in metabolic assessment of cultured cells and perfused tissues. Methods Enzymol. 2015, 561, 73−106. (16) Khemtong, C.; Carpenter, N. R.; Lumata, L. L.; Merritt, M. E.; Moreno, K. X.; Kovacs, Z.; Malloy, C. R.; Sherry, A. D. Hyperpolarized 13C NMR detects rapid drug-induced changes in cardiac metabolism. Magn. Reson. Med. 2015, 74, 312−319. (17) Jensen, P. R.; Peitersen, T.; Karlsson, M.; Zandt, R. I.; Gisselsson, A.; Hansson, G.; Meier, S.; Lerche, M. H. Tissue-specific Short Chain Fatty Acid Metabolism and Slow Metabolic Recovery after Ischemia from Hyperpolarized NMR in Vivo. J. Biol. Chem. 2009, 284, 36077−36082. (18) Schroeder, M. A.; Cochlin, L. E.; Heather, L. C.; Clarke, K.; Radda, G. K.; Tyler, D. J. In vivo assessment of pyruvate dehydrogenase flux in the heart using hyperpolarized carbon-13 magnetic resonance. Proc. Natl. Acad. Sci. U. S. A. 2008, 105, 12051− 12056. (19) Meier, S.; Jensen, P. R.; Duus, J. O. Real-time detection of central carbon metabolism in living Escherichia coli and its response to perturbations. FEBS Lett. 2011, 585, 3133−3138. (20) Schroeder, M. A.; Atherton, H. J.; Ball, D. R.; Cole, M. A.; Heather, L. C.; Griffin, J. J.; Clarke, K.; Radda, G. K.; Tyler, D. J. Realtime assessment of Krebs cycle metabolism using hyperpolarized 13C magnetic resonance spectroscopy. FASEB J. 2009, 23, 2529−2538. (21) Gallagher, F. A.; Kettunen, M. I.; Day, S. E.; Hu, D. E.; Ardenkjær-Larsen, J. H.; ‘t Zandt, R. I.; Jensen, P. R.; Karlsson, M.; Golman, K.; Lerche, M. H.; Brindle, K. M. Magnetic resonance imaging of pH in vivo using hyperpolarized 13C-labelled bicarbonate. Nature 2008, 453, 940−943. (22) Schroeder, M. A.; Atherton, H. J.; Cochlin, L. E.; Clarke, K.; Radda, G. K.; Tyler, D. J. The effect of hyperpolarized tracer concentration on myocardial uptake and metabolism. Magn. Reson. Med. 2009, 61, 1007−1014. (23) Gallagher, F. A.; Kettunen, M. I.; Day, S. E.; Hu, D. E.; Karlsson, M.; Gisselsson, A.; Lerche, M. H.; Brindle, K. M. Detection of tumor glutamate metabolism in vivo using 13C magnetic resonance spectroscopy and hyperpolarized [1-13C]glutamate. Magn. Reson. Med. 2011, 66, 18−23. (24) Schroeder, M. A.; Atherton, H. J.; Cochlin, L. E.; Clarke, K.; Radda, G. K.; Tyler, D. J. The effect of hyperpolarized tracer concentration on myocardial uptake and metabolism. Magn. Reson. Med. 2009, 61, 1007−1014. (25) Karlsson, M.; Jensen, P. R.; Zandt, R. I.; Gisselsson, A.; Hansson, G.; Duus, J. O.; Meier, S.; Lerche, M. H. Imaging of branched chain amino acid metabolism in tumors with hyperpolarized 13 C ketoisocaproate. Int. J. Cancer 2010, 127, 729−736. (26) Lumata, L.; Ratnakar, S. J.; Jindal, A.; Merritt, M.; Comment, A.; Malloy, C.; Sherry, A. D.; Kovacs, Z. BDPA: An Efficient Polarizing Agent for Fast Dissolution Dynamic Nuclear Polarization NMR Spectroscopy. Chem. - Eur. J. 2011, 17, 10825−10827.

(27) Lumata, L.; Merritt, M.; Malloy, C.; Sherry, A. D.; van Tol, J.; Song, L.; Kovacs, Z. Dissolution DNP-NMR spectroscopy using galvinoxyl as a polarizing agent. J. Magn. Reson. 2013, 227, 14−19. (28) Lumata, L.; Merritt, M.; Khemtong, C.; Ratnakar, S. J.; van Tol, J.; Yu, L.; Song, L.; Kovacs, Z. The efficiency of DPPH as a polarising agent for DNP-NMR spectroscopy. RSC Adv. 2012, 2, 12812−12817. (29) Niedbalski, P.; Parish, C.; Kiswandhi, A.; Lumata, L. 13C dynamic nuclear polarization using isotopically-enriched 4-oxoTEMPO free radicals. Magn. Reson. Chem. 2016, 54, 962−967. (30) Lumata, L.; Kovacs, Z.; Sherry, A. D.; Malloy, C.; Hill, S.; van Tol, J.; Yu, L.; Song, L.; Merritt, M. E. Electron Spin Resonance Studies of Trityl OX063 at a Concentration Optimal for DNP. Phys. Chem. Chem. Phys. 2013, 15, 9800−9807. (31) Heckmann, J.; Meyer, W.; Radtke, E.; Reicherz, G.; Goertz, S. Electron Spin Resonance and Its Implication on the Maximum Nuclear Polarization of Deuterated Solid Target Materials. Phys. Rev. B: Condens. Matter Mater. Phys. 2006, 74, 134418. (32) Yoshihara, H. A. I.; Can, E.; Karlsson, M.; Lerche, M. H.; Schwitter, J.; Comment, A. High-field dissolution dynamic nuclear polarization of [1-13C]pyruvic acid. Phys. Chem. Chem. Phys. 2016, 18, 12409−12413. (33) Ardenkjaer-Larsen, J. H.; Macholl, S.; Jóhannesson, H. Dynamic Nuclear Polarization with Trityls at 1.2 K. Appl. Magn. Reson. 2008, 34, 509−522. (34) Lumata, L.; Merritt, M. E.; Malloy, C. R.; Sherry, A. D.; Kovacs, Z. Impact of Gd3+ on DNP of [1-13C]Pyruvate Doped with Trityl OX063, BDPA, or 4-oxo-TEMPO. J. Phys. Chem. A 2012, 116, 5129− 5138. (35) Kiswandhi, A.; Lama, B.; Niedbalski, P.; Goderya, M.; Long, J.; Lumata, L. RSC Adv. 2016, 6, 38855−38860. (36) Lumata, L. L.; Martin, R.; Jindal, A. K.; Kovacs, Z.; Conradi, M. S.; Merritt, M. E. Development and performance of a 129 GHz dynamic nuclear polarizer in an ultra-wide bore superconducting magnet. MAGMA 2015, 28, 195−205. (37) Kiswandhi, A.; Niedbalski, P.; Parish, C.; Ferguson, S.; Taylor, D.; McDonald, G.; Lumata, L. Construction and 13C hyperpolarization efficiency of a 180 GHz dissolution dynamic nuclear polarization system. Magn. Reson. Chem. 2017, DOI: 10.1002/ mrc.4597. (38) Comment, A.; van den Brandt, B.; Uffmann, K.; Kurdzesau, F.; Jannin, S.; Konter, J. A.; Hautle, P.; Wenckebach, W. T.; Gruetter, R.; van der Klink, J. J. Design and performance of a DNP prepolarizer coupled to a rodent MRI scanner. Concepts Magn. Reson., Part B 2007, 31B, 255−269. (39) Kurdzesau, F.; van den Brandt, B.; Comment, A.; Hautle, P.; Jannin, S.; van der Klink, J. J.; Konter, J. A. Dynamic Nuclear Polarization of Small Labeled Molecules in Frozen Water−alcohol Solutions. J. Phys. D: Appl. Phys. 2008, 41, 155506. (40) Batel, M.; Krajewski, M.; Däpp, A.; Hunkeler, A.; Meier, B. H.; Kozerke, S.; Ernst, M. Dissolution dynamic nuclear polarization efficiency enhanced by Hartmann−Hahn cross polarization. Chem. Phys. Lett. 2012, 554, 72−76. (41) Niedbalski, P.; Parish, C.; Kiswandhi, A.; Kovacs, Z.; Lumata, L. Influence of 13C Isotopic Labeling Location on Dynamic Nuclear Polarization of Acetate. J. Phys. Chem. A 2017, 121, 3227−3233. (42) Lumata, L.; Kovacs, Z.; Malloy, C.; Sherry, A. D.; Merritt, M. Effect of 13C Enrichment in the Glassing Matrix on Dynamic Nuclear Polarization of [1-13C]pyruvate. Phys. Med. Biol. 2011, 56, N85−N92. (43) Lumata, L.; Merritt, M. E.; Kovacs, Z. Influence of Deuteration in the Glassing Matrix on 13C Dynamic Nuclear Polarization. Phys. Chem. Chem. Phys. 2013, 15, 7032−7035. (44) Friesen-Waldner, L.; Chen, A.; Mander, W.; Scholl, T. J.; McKenzie, C. A. Optimisation of Dynamic Nuclear Polarisation of [1-13C] Pyruvate by Addition of Gadolinium-Based Contrast Agents. J. Magn. Reson. 2012, 223, 85−89. (45) Gordon, J. W.; Fain, S. B.; Rowland, I. J. Effect of Lanthanide Ions on Dynamic Nuclear Polarization Enhancement and Liquid-State T1 Relaxation. Magn. Reson. Med. 2012, 68, 1949−1954. H

DOI: 10.1021/acs.jpca.7b03869 J. Phys. Chem. A XXXX, XXX, XXX−XXX

Article

The Journal of Physical Chemistry A

Mechanisms in Solid State 13C DNP at 95 GHz Using Trityl Radicals. J. Magn. Reson. 2013, 230, 212−219. (67) Ravera, E.; Shimon, D.; Feintuch, A.; Goldfarb, D.; Vega, S.; Flori, A.; Luchinat, C.; Menichetti, L.; Parigi, G. The Effect of Gd on Trityl-Based Dynamic Nuclear Polarisation in Solids. Phys. Chem. Chem. Phys. 2015, 17, 26969−26978. (68) Milani, J.; Vuichoud, B.; Bornet, A.; Miéville, P.; Mottier, R.; Jannin, S.; Bodenhausen, G. A magnetic tunnel to shelter hyperpolarized fluids. Rev. Sci. Instrum. 2015, 86, 024101. (69) Gries, H. Extracellular MRI Contrast Agents Based on Gadolinium. In Contrast Agents I; Krause, P. D. W., Ed.; Topics in Current Chemistry; Springer: Berlin, Heidelberg, 2002; pp 1−24. (70) Caravan, P. Strategies for Increasing the Sensitivity of Gadolinium Based MRI Contrast Agents. Chem. Soc. Rev. 2006, 35, 512−523. (71) Song, Y.; Kohlmeir, E. K.; Meade, T. J. Synthesis of Multimeric MR Contrast Agents for Cellular Imaging. J. Am. Chem. Soc. 2008, 130, 6662−6663.

(46) Kiswandhi, A.; Niedbalski, P.; Parish, C.; Kaur, P.; Martins, A.; Fidelino, L.; Khemtong, C.; Song, L.; Sherry, A. D.; Lumata, L. Impact of Ho3+-doping on 13C dynamic nuclear polarization using trityl OX063 free radical. Phys. Chem. Chem. Phys. 2016, 18, 21351−21359. (47) Niedbalski, P.; Parish, C.; Kiswandhi, A.; Fidelino, L.; Khemtong, C.; Hayati, Z.; Song, L.; Martins, A.; Sherry, A. D.; Lumata, L. Influence of Dy3+ and Tb3+-doping on 13C dynamic nuclear polarization. J. Chem. Phys. 2017, 146, 014303. (48) Aime, S.; Botta, M.; Ermondi, G. NMR Study of Solution Structures and Dynamics of lanthanide(III) Complexes of DOTA. Inorg. Chem. 1992, 31, 4291−4299. (49) Wang, X.; Jin, T.; Comblin, V.; Lopez-Mut, A.; Merciny, E.; Desreux, J. F. A Kinetic Investigation of the Lanthanide DOTA Chelates. Stability and Rates of Formation and of Dissociation of a Macrocyclic gadolinium(III) Polyaza Polycarboxylic MRI Contrast Agent. Inorg. Chem. 1992, 31, 1095−1099. (50) Patyal, B. R.; Gao, J.-H.; Williams, R. F.; Roby, J.; Saam, B.; Rockwell, B. A.; Thomas, R. J.; Stolarski, D. J.; Fox, P. T. Longitudinal Relaxation and Diffusion Measurements Using Magnetic Resonance Signals From Laser-Hyperpolarized 129Xe Nuclei. J. Magn. Reson. 1997, 126, 58−65. (51) Wolber, J.; Ellner, F.; Fridlund, B.; Gram, A.; Jóhannesson, H.; Hansson, G.; Hansson, L. H.; Lerche, M. H.; Månsson, S.; Servin, R.; et al. Generating Highly Polarized Nuclear Spins in Solution Using Dynamic Nuclear Polarization. Nucl. Instrum. Methods Phys. Res., Sect. A 2004, 526, 173−181. (52) Reynolds, S.; Patel, H. Monitoring the Solid-State Polarization of 13C, 15N, 2H, 29Si and 31P. Appl. Magn. Reson. 2008, 34, 495−508. (53) Goertz, S. T. The Dynamic Nuclear Polarization Process. Nucl. Instrum. Methods Phys. Res., Sect. A 2004, 526, 28−42. (54) Borghini, M. Spin-Temperature Model of Nuclear Dynamic Polarization Using Free Radicals. Phys. Rev. Lett. 1968, 20, 419−421. (55) Wenckebach, W. T. The Solid Effect. Appl. Magn. Reson. 2008, 34, 227−235. (56) Hovav, Y.; Feintuch, A.; Vega, S. Theoretical Aspects of Dynamic Nuclear Polarization in the Solid State − The Cross Effect. J. Magn. Reson. 2012, 214, 29−41. (57) Mance, D.; Gast, P.; Huber, M.; Baldus, M.; Ivanov, K. L. The Magnetic Field Dependence of Cross-Effect Dynamic Nuclear Polarization under Magic Angle Spinning. J. Chem. Phys. 2015, 142, 234201. (58) Thurber, K. R.; Tycko, R. Theory for Cross Effect Dynamic Nuclear Polarization under Magic-Angle Spinning in Solid State Nuclear Magnetic Resonance: The Importance of Level Crossings. J. Chem. Phys. 2012, 137, 084508. (59) Walker, S. A.; Edwards, D. T.; Siaw, T. A.; Armstrong, B. D.; Han, S. Temperature Dependence of High Field 13C Dynamic Nuclear Polarization Processes with Trityl Radicals below 35 K. Phys. Chem. Chem. Phys. 2013, 15, 15106−15120. (60) Abragam, A.; Bleaney, B. Electron paramagnetic resonance of transition ions; Clarendon Press: Oxford, 1970. (61) Jannin, S.; Comment, A.; van der Klink, J. J. Dynamic Nuclear Polarization by Thermal Mixing Under Partial Saturation. Appl. Magn. Reson. 2012, 43, 59−68. (62) Wenckebach, W. T. Dynamic nuclear polarization via thermal mixing: Beyond the high temperature approximation. J. Magn. Reson. 2017, 277, 68−78. (63) Serra, S. C.; Rosso, A.; Tedoldi, F. On the Role of Electron− nucleus Contact and Microwave Saturation in Thermal Mixing DNP. Phys. Chem. Chem. Phys. 2013, 15, 8416−8428. (64) Colombo Serra, S.; Filibian, M.; Carretta, P.; Rosso, A.; Tedoldi, F. Relevance of Electron Spin Dissipative Processes to Dynamic Nuclear Polarization via Thermal Mixing. Phys. Chem. Chem. Phys. 2014, 16, 753−764. (65) Serra, S. C.; Rosso, A.; Tedoldi, F. Electron and Nuclear Spin Dynamics in the Thermal Mixing Model of Dynamic Nuclear Polarization. Phys. Chem. Chem. Phys. 2012, 14, 13299−13308. (66) Banerjee, D.; Shimon, D.; Feintuch, A.; Vega, S.; Goldfarb, D. The Interplay between the Solid Effect and the Cross Effect I

DOI: 10.1021/acs.jpca.7b03869 J. Phys. Chem. A XXXX, XXX, XXX−XXX