Wednesday, October 31, 2012

Nuclear Magnetic Resonance of Hyperpolarized Fluorine for Characterization of Protein–Ligand Interactions

Lee, Y., et al., Nuclear Magnetic Resonance of Hyperpolarized Fluorine for Characterization of Protein–Ligand Interactions. J. Am. Chem. Soc., 2012. 134(42): p. 17448-17451.

Fluorine NMR spectroscopy is widely used for detection of protein?ligand interactions in drug discovery because of the simplicity of fluorine spectra combined with a relatively high likelihood for a drug molecule to include at least one fluorine atom. In general, an important limitation of NMR spectroscopy in drug discovery is its sensitivity, which results in the need for unphysiologically high protein concentrations and large ligand:protein ratios. An enhancement in the 19F signal of several thousand fold by dynamic nuclear polarization allows for the detection of submicromolar concentrations of fluorinated small molecules. Techniques for exploiting this gain in signal to detect ligands in the strong-, intermediate-, and weak-binding regimes are presented. Similar to conventional NMR analysis, dissociation constants are determined. However, the ability to use a low ligand concentration permits the detection of ligands in slow exchange that are not easily amenable to drug screening by traditional NMR methods. The relative speed and additional information gained may make the hyperpolarization-based approach an interesting alternative for use in drug discovery.

Friday, October 26, 2012

Fast passage dynamic nuclear polarization on rotating solids

Mentink-Vigier, F., et al., Fast passage dynamic nuclear polarization on rotating solids. J. Magn. Reson., 2012. 224(0): p. 13-21.

Magic Angle Spinning (MAS) Dynamic Nuclear Polarization (DNP) has proven to be a very powerful way to improve the signal to noise ratio of NMR experiments on solids. The experiments have in general been interpreted considering the Solid-Effect (SE) and Cross-Effect (CE) DNP mechanisms while ignoring the influence of sample spinning. In this paper, we show experimental data of MAS-DNP enhancements of 1H and 13C in proline and SH3 protein in glass forming water/glycerol solvent containing TOTAPOL. We also introduce a theoretical model that aims at explaining how the nuclear polarization is built in MAS-DNP experiments. By using Liouville space based simulations to include relaxation on two simple spin models, {electron–nucleus} and {electron–electron–nucleus}, we explain how the basic MAS-SE-DNP and MAS-CE-DNP processes work. The importance of fast energy passages and short level anti-crossing is emphasized and the differences between static DNP and MAS-DNP is explained. During a single rotor cycle the enhancement in the {electron–electron–nucleus} system arises from MAS-CE-DNP involving at least three kinds of two-level fast passages: an electron–electron dipolar anti-crossing, a single quantum electron MW encounter and an anti-crossing at the CE condition inducing nuclear polarization in- or decrements. Numerical, powder-averaged, simulations were performed in order to check the influence of the experimental parameters on the enhancement efficiencies. In particular we show that the spinning frequency dependence of the theoretical MAS-CE-DNP enhancement compares favorably with the experimental 1H and 13C MAS-DNP enhancements of proline and SH3.

Thursday, October 25, 2012

4th International DNP Symposium in Copenhagen

Dear Friends and Colleagues 

We are happy to announce the "4th International DNP Symposium", Aug 28-31, 2013, in Copenhagen, Denmark: Please mark your calenders. 

The meeting follows the highly successful previous meetings in Nottingham (2007), Koenigstein (2009) and Lausanne (2011). 

Over the three days of the meeting there will be series of talks from esteemed researchers in the field assessing the present state of the art. The symposium will cover the basic science across physics, chemistry and biology as well as the engineering challenges and applications. DNP and hyperpolarization has lead to some very significant new applications in NMR and MRI, and we are only at the beginning of exploring the opportunities. 

The symposium will accept abstracts for oral and poster presentation selection. 

Registration will open January 2013, and we will send you a reminder email as soon as the website is open for registration. 

The organizing committee, 

Jan H. Ardenkjaer-Larsen 

Lars G. Hanson 
Charlotte H Gotfredsen 
Susanne L. Mossin 
Sebastian Meier 
Lise V. Søgaard 
Niels Chr Nielsen 

Monday, October 22, 2012

Transfer of the Haupt-hyperpolarization to neighbor spins

Icker, M., P. Fricke, and S. Berger, Transfer of the Haupt-hyperpolarization to neighbor spins. J. Magn. Reson., 2012. 223(0): p. 148-150.

The NMR hyperpolarization observed for freely rotating methyl groups by exerting a temperature jump from 4.2 K to 298 K can be transferred to spins which have a spin, spin coupling with the carbon of the methyl group. First, a spin echo sequence readjusts the primary up/down signals to an in-phase multiplet. This in-phase magnetization is then decoupled and transferred by a simple COSY step using one scan. The polarization factors at the neighbor spins are about 50 by comparing their signal-to-noise ratio with the signal strength after full relaxation.

Friday, October 19, 2012

Optimisation of dynamic nuclear polarisation of [1-13C] pyruvate by addition of gadolinium-based contrast agents

Friesen-Waldner, L., et al., Optimisation of dynamic nuclear polarisation of [1-13C] pyruvate by addition of gadolinium-based contrast agents. J. Magn. Reson., 2012. 223(0): p. 85-89.

Dynamic nuclear polarisation (DNP) of carbon-13 (13C) enriched endogenous compounds provides a novel means for magnetic resonance imaging and spectroscopy of biological processes. Adding small amounts of gadolinium-based contrast agents (GBCAs) to the 13C-enriched substrate matrix increases the amount of hyperpolarisation that can be achieved, but also may decrease the longitudinal relaxation time (T1) of the 13C nucleus in solution. This study examined the effects of five different GBCA at concentrations of 0.5, 1, 2, and 3 mM on [1-13C]-enriched pyruvic acid. It was found that contrast agents with an open chain structure (Gadobenate dimeglumine, Gadopentetate dimeglumine, Gadodiamide) caused the largest enhancement (up to 82%) in solid state polarisation relative to solutions without GBCA. In the liquid state, T1 of pyruvate decreased by as much as 62% and polarisation was much lower (70%) relative to solutions without GBCA added. Conversely, for GBCA with macrocyclic structures (Gadoterate meglumine, Gadoteridol), the solid state polarisation enhancement was only slightly less than the open chain GBCA, but enhanced polarisation was retained much better in the liquid state with minimal decrease in T1 (25% at the highest GBCA concentrations). Near maximum polarisation in the solid state was obtained at a GBCA concentration of 2 mM, with a higher concentration of 3 mM producing minimal improvement. These results indicate that the macrocyclic contrast agents provide the best combination of high solid state and liquid state polarisations with minimal loss of T1 in experiments with hyperpolarised 13C-enriched pyruvate. This suggests that macrocyclic contrast agents should be the GBCA of choice for maximising signal in experiments with hyperpolarised 13C-enriched pyruvate, particularly for in vivo measurements where shortened substrate T1 is especially problematic.

Tuesday, October 16, 2012

Simple and Highly Sensitive Measurement Method for Detection of Glass Transition Temperatures of Polymers: Application of ESR Power Saturation Phenomenon with Conventional Spin-Probe Technique

This is not an article about DNP spectroscopy. However, it shows how EPR spectroscopy and nitroxide spin labels can be used to characterize mobility of soft matter through shape analysis of the EPR spectrum. Since mobility and relaxation is directly connected I thought this article could also be of interest to the DNP community.

Miwa, Y. and K. Yamamoto, Simple and Highly Sensitive Measurement Method for Detection of Glass Transition Temperatures of Polymers: Application of ESR Power Saturation Phenomenon with Conventional Spin-Probe Technique. The Journal of Physical Chemistry B, 2012. 116(30): p. 9277-9284.

A combination of the microwave power saturation (MPS) method of electron spin resonance (ESR) and spin probing is proposed as a simple and practical technique for detecting the glass transition temperatures, Tg, of polymers with high sensitivity. Effects of the spin-probe size and concentration on the Tg value of polystyrene (PS) determined by MPS, Tg,ESR, were first evaluated. Spin-probed PS with four types of nitroxides, namely, di-tert-butyl nitroxide (DBN), 2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO), 4-benzoyloxy-2,2,6,6-tetramethylpiperidine-1-oxyl (BZONO), and 4?,4?-dimethyl-spiro(5α-cholestane-3,2?-oxazolidin)-3?-yloxy free radical (CHOL), having molecular weights of 144, 156, 276, and 473, respectively, and spin-labeled PS with TEMPO were prepared. The Tg,ESR values for the spin-probed PS with DBN, TEMPO, BZONO, and CHOL and spin-labeled PS were determined to 360, 363, 374, 374, and 375 K, respectively, within experimental uncertainties of 2 K, whereas the glass transition temperature determined by DSC, Tg,DSC, was 375 K for all samples. A significant decrease in Tg,ESR for small spin probes was shown to be due to decoupling between the mobilities of small spin probes and PS segments. Concerning the concentration, a decrease in the saturation factor, S, induced by shortening of the spin?spin relaxation time was observed for the spin-probed PS with CHOL when the concentration of CHOL was more than 1.0 wt %. Furthermore, Tg,ESR decreased slightly with increasing weight fraction of CHOL because of the ?plasticizer effect? of CHOL. However, the Tg,ESR and Tg,DSC values corresponded for each concentration. Thus, large spin probes, such as CHOL and BZONO, are appropriate for the determination of Tg,ESR values; the concentration of the spin probes does not affect the Tg,ESR value unless the overall Tg value is reduced by blending of excess spin probes. Finally, measurements of Tg,ESR in PS/silica composites containing more than 95 wt % silica are shown as an application example of the present method. Tg,ESR was clearly determined even for the PS/silica composites with 98 wt % silica; a decrease in Tg,ESR with increasing silica content was observed.

Monday, October 15, 2012

Distance measurements across randomly distributed nitroxide probes from the temperature dependence of the electron spin phase memory time at 240GHz

Edwards, D.T., et al., Distance measurements across randomly distributed nitroxide probes from the temperature dependence of the electron spin phase memory time at 240GHz. J. Magn. Reson., 2012. 223(0): p. 198-206.

At 8.5 T, the polarization of an ensemble of electron spins is essentially 100% at 2 K, and decreases to 30% at 20 K. The strong temperature dependence of the electron spin polarization between 2 and 20 K leads to the phenomenon of spin bath quenching: temporal fluctuations of the dipolar magnetic fields associated with the energy-conserving spin “flip-flop” process are quenched as the temperature of the spin bath is lowered to the point of nearly complete spin polarization. This work uses pulsed electron paramagnetic resonance (EPR) at 240 GHz to investigate the effects of spin bath quenching on the phase memory times (TM) of randomly-distributed ensembles of nitroxide molecules below 20 K at 8.5 T. For a given electron spin concentration, a characteristic, dipolar flip-flop rate (W) is extracted by fitting the temperature dependence of TM to a simple model of decoherence driven by the spin flip-flop process. In frozen solutions of 4-Amino-TEMPO, a stable nitroxide radical in a deuterated water–glass, a calibration is used to quantify average spin–spin distances as large as <r> = 6.6 nm from the dipolar flip-flop rate. For longer distances, nuclear spin fluctuations, which are not frozen out, begin to dominate over the electron spin flip-flop processes, placing an effective ceiling on this method for nitroxide molecules. For a bulk solution with a three-dimensional distribution of nitroxide molecules at concentration n, we find W ∝ n ∝ 1 / r ¯ 3 , which is consistent with magnetic dipolar spin interactions. Alternatively, we observe W ∝ n 3 2 for nitroxides tethered to a quasi two-dimensional surface of large (Ø ∼ 200 nm), unilamellar, lipid vesicles, demonstrating that the quantification of spin bath quenching can also be used to discern the geometry of molecular assembly or organization.

Friday, October 12, 2012

A 140 GHz pulsed EPR/212 MHz NMR spectrometer for DNP studies

Smith, A.A., et al., A 140 GHz pulsed EPR/212 MHz NMR spectrometer for DNP studies. J. Magn. Reson., 2012. 223(0): p. 170-179.

We described a versatile spectrometer designed for the study of dynamic nuclear polarization (DNP) at low temperatures and high fields. The instrument functions both as an NMR spectrometer operating at 212 MHz (1H frequency) with DNP capabilities, and as a pulsed-EPR operating at 140 GHz. A coiled TE011 resonator acts as both an NMR coil and microwave resonator, and a double balanced (1H, 13C) radio frequency circuit greatly stabilizes the NMR performance. A new 140 GHz microwave bridge has also been developed, which utilizes a four-phase network and ELDOR channel at 8.75 GHz, that is then multiplied and mixed to obtain 140 GHz microwave pulses with an output power of 120 mW. Nutation frequencies obtained are as follows: 6 MHz on S = 1/2 electron spins, 100 kHz on 1H, and 50 kHz on 13C. We demonstrate basic EPR, ELDOR, ENDOR, and DNP experiments here. Our solid effect DNP results demonstrate an enhancement of 144 and sensitivity gain of 310 using OX063 trityl at 80 K and an enhancement of 157 and maximum sensitivity gain of 234 using Gd-DOTA at 20 K, which is significantly better performance than previously reported at high fields (⩾3 T).

Wednesday, October 10, 2012

First determination of the spin relaxation properties of a nitronyl nitroxide in solution by electron spin echoes at X-band: A comparison with Tempone

This article describes no DNP results. However, it can be still of interest for the solution-state DNP community since it discusses relaxation times of nitroxide spin-labels.

Collauto, A., A. Barbon, and M. Brustolon, First determination of the spin relaxation properties of a nitronyl nitroxide in solution by electron spin echoes at X-band: A comparison with Tempone. J. Magn. Reson., 2012. 223(0): p. 180-186.

We studied by electron spin echo pulse methods the spin relaxation properties of a phenyl nitronyl nitroxide radical (2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide, PTIO) at X-band in fluid toluene solution in a wide temperature range, and in a water/glycerol 1:1 mixture near room temperature. The relaxation properties of PTIO have been compared with that of Tempone, as a widely used nitroxide. By a new procedure, based on experimental results on the temperature dependences of the relaxation times T1 and T2, and on the approximation of an isotropic Brownian rotational diffusion, we separated non-secular, spin rotational and residual terms from the transverse relaxation rate to isolate secular and pseudosecular contributions. By comparing the results for the two radicals we found the differences in the magnetic properties that give rise to slower transverse (T2) and longitudinal (T1) electron spin relaxation for PTIO in the whole temperature range explored in this work.

Monday, October 8, 2012

Field dependence of the relaxation of 79Br in KBr and its use as a temperature calibrant

Beckett, P., et al., Field dependence of the relaxation of 79Br in KBr and its use as a temperature calibrant. J. Magn. Reson., 2012. 223(0): p. 61-63.

The longitudinal relaxation time of 79Br nuclei in KBr is field independent, between 4.7 T and 14.1 T. The results suggest that inconsistencies in the literature are due to differences in the experimental set-ups. The limitations of KBr as temperature calibrant are discussed.

Friday, October 5, 2012

Dynamic Nuclear Polarization NMR Spectroscopy of Microcrystalline Solids

This is a very interesting article, since it demonstrates how DNP can be tremendously helpful for the characterization of small molecules (e.g. quality control of pharmaceutical formulas). Here, DNP-enhanced 13C-13C correlation spectra are presented that are recorded using unlabeled material and the glass-forming solvents were carefully chosen so the analyte remains unaltered.

Rossini, A.J., et al., Dynamic Nuclear Polarization NMR Spectroscopy of Microcrystalline Solids. J. Am. Chem. Soc., 2012.

Dynamic nuclear polarization (DNP) solid-state NMR has been applied to powdered microcrystalline solids to obtain sensitivity enhancements on the order of 100. Glucose, sulfathiazole, and paracetamol were impregnated with bis-nitroxide biradical (bis-cyclohexyl-TEMPO-bisketal, bCTbK) solutions of organic solvents. The organic solvents were carefully chosen to be nonsolvents for the compounds, so that DNP-enhanced solid-state NMR spectra of the unaltered solids could be acquired. A theoretical model is presented that illustrates that for externally doped organic solids characterized by long spin?lattice relaxation times (T1(1H) > 200 s), 1H?1H spin diffusion can relay enhanced polarization over micrometer length scales yielding substantial DNP enhancements (ε). ε on the order of 60 are obtained for microcrystalline glucose and sulfathiazole at 9.4 T and with temperatures of ca. 105 K. The large gain in sensitivity enables the rapid acquisition of 13C-13C correlation spectra at natural isotopic abundance. It is anticipated that this will be a general method for enhancing the sensitivity of solid-state NMR experiments of organic solids.

Wednesday, October 3, 2012

Techniques in DNP-NMR Spectroscopy

Check out my second blog post that I wrote for the SpinSights blog. It gives a short overview about the different techniques of DNP-NMR spectroscopy.

Here is a quick summary:

  • DNP can be separated into four different areas based on whether DNP is performed at high or low magnetic field strengths (1,2 vs. 3,4) or in the solid or solution state (1,3 vs. 2,4). In principle it is always possible to directly polarize a sample, but some (direct) polarizing methods are more challenging than others. E.g. at low magnetic fields (cases 3 and 4) a solid-state source can be used, but the output power of such solid-sate microwave/THz sources drops off drastically at higher frequencies. In these cases a gyrotron can be used (cases 1 and 2).
  • For solution-state NMR the situation is more challenging due to the high ohmic losses of the liquid NMR sample and severe heating that occurs (think microwave oven). To prevent this, a resonance structure (resonator) is required to separate the electric field components of the THz radiation from the magnetic field components. This becomes very challenging since at high frequencies (e.g. 400 MHz 1H, 268 GHz e-) the wavelength of the THz radiation is 1.12 mm and therefore fundamental mode devices are very small and can accommodate only very small amounts of liquid sample. This has led to the development of several other techniques:
    • Temperature-Jump DNP (A): In a TJ-DNP experiment the sample (10 – 20 microliters) is polarized in-situ at 90 K using a gyrotron as the THz source. Once the sample is polarized, the sample is rapidly melted using a laser flash, and the NMR experiment can be performed. After the laser is turned off, the sample freezes and can be polarized again.
    • Dissolution-DNP (B): In a dissolution experiment the sample is polarized ex-situ in the solid-state at liquid Helium temperatures at 3.3T using a low-power solid-state 95 GHz microwave source. The sample is then melted using a hot solvent and rapidly transferred into the high-field NMR spectrometer for detection.
    • Shuttle DNP9-10 (arrow C): In a shuttle-DNP experiment, the sample is polarized at a low magnetic field and then physically shuttled into the high-field NMR magnet to acquire the NMR spectrum.

Monday, October 1, 2012

Physical methods and techniques NMR spectroscopy

This article came a bit as a surprise. I didn't expect DNP to show up in a review article published in a journal for organic chemistry. However, I'm very delighted to see that since it shows that researchers from many different areas become aware of the method.

Edgar, M., Physical methods and techniques NMR spectroscopy. Annual Reports Section "B" (Organic Chemistry), 2012. 108(0): p. 292-315.

NMR spectroscopy continues to evolve, with publications in 2011 providing an eclectic collection of applications, advances and incremental improvements. Publications highlights include: DNP in liquids and solids, decoupling effects at high spinning speed in solid-state, results from the 80 kHz spinning-speed 1 mm MAS rotor, enhanced diffusion methods, results from the new 1.0 GHz magnet, an intent to create a 1.3 GHz magnet, results from the 2.6 GHz pulsed-magnet, a cryogen-free MRI, and the use of multiple receivers to acquire multiple experiments within the same pulse sequence. I am indebted to the great and the good whose work has been reported here, all credit is theirs; errors, omissions and blame are all mine.