Category Archives: Small Molecules

Real‐Time Interrogation of Aspirin Reactivity, Biochemistry, and Biodistribution by Hyperpolarized Magnetic Resonance Spectroscopy

Zacharias, Niki M., Argentina Ornelas, Jaehyuk Lee, Jingzhe Hu, Jennifer S. Davis, Nasir Uddin, Shivanand Pudakalakatti, et al. “Real‐Time Interrogation of Aspirin Reactivity, Biochemistry, and Biodistribution by Hyperpolarized Magnetic Resonance Spectroscopy.” Angewandte Chemie 131, no. 13 (March 22, 2019): 4223–27.

https://doi.org/10.1002/ange.201812759

Hyperpolarized magnetic resonance spectroscopy (HPMR) enables quantitative, non-radioactive, real-time measurement of imaging probe biodistribution and metabolism in vivo. Here, we investigate and report on the development and characterization of hyperpolarized acetylsalicylic acid (aspirin) and its use as a nuclear magnetic resonance (NMR) probe. Aspirin derivatives were synthesized with single- and double-13C labels and hyperpolarized by dynamic nuclear polarization with 4.7% and 3% polarization, respectively. The longitudinal relaxation constants (T1) for the labeled acetyl and carboxyl carbonyls were approximately 30 seconds, supporting in vivo imaging and spectroscopy applications. In vitro hydrolysis, transacetylation, and albumin binding of hyperpolarized aspirin were readily monitored in real time by 13C-NMR spectroscopy. Hyperpolarized, double-labeled aspirin was well tolerated in mice and could be observed by both 13C-MR imaging and 13C-NMR spectroscopy in vivo.

Polymorphs of Theophylline Characterized by DNP Enhanced Solid-State NMR

Pinon, A.C., et al., Polymorphs of Theophylline Characterized by DNP Enhanced Solid-State NMR. Mol Pharm, 2015.

http://www.ncbi.nlm.nih.gov/pubmed/26393368

We show how dynamic nuclear polarization (DNP) enhanced solid-state NMR spectroscopy can be used to characterize polymorphs and solvates of organic solids. We applied DNP to three polymorphs and one hydrated form of the asthma drug molecule theophylline. For some forms of theophylline, sample grinding and impregnation with the radical-containing solution, which are necessary to prepare the samples for DNP, were found to induce polymorphic transitions or desolvation between some forms. We present protocols for sample preparation for solid-state magic-angle spinning (MAS) DNP experiments that avoid the polymorphic phase transitions in theophylline. These protocols include cryogrinding, grinding under inert atmosphere, and the appropriate choice of the impregnating liquid. By applying these procedures, we subsequently demonstrate that two-dimensional correlation experiments, such as 1H-13C and 1H-15N HETCOR or 13C-13C INADEQUATE, can be obtained at natural isotopic abundance in reasonable times, thus enabling more advanced structural characterization of polymorphs.

High-resolution NMR of hydrogen in organic solids by DNP enhanced natural abundance deuterium spectroscopy

Rossini, A.J., et al., High-resolution NMR of hydrogen in organic solids by DNP enhanced natural abundance deuterium spectroscopy. J. Magn. Reson., 2015. 259: p. 192-198.

http://dx.doi.org/10.1016/j.jmr.2015.08.020

We demonstrate that high field (9.4 T) dynamic nuclear polarization (DNP) at cryogenic (∼100 K) sample temperatures enables the rapid acquisition of natural abundance 1H–2H cross-polarization magic angle spinning (CPMAS) solid-state NMR spectra of organic solids. Spectra were obtained by impregnating substrates with a solution of the stable DNP polarizing agent TEKPol in tetrachloroethane. Tetrachloroethane is a non-solvent for the solids, and the unmodified substrates are then polarized through spin diffusion. High quality natural abundance 2H CPMAS spectra of histidine hydrochloride monohydrate, glycylglycine and theophylline were acquired in less than 2 h, providing direct access to hydrogen chemical shifts and quadrupolar couplings. The spectral resolution of the 2H solid-state NMR spectra is comparable to that of 1H spectra obtained with state of the art homonuclear decoupling techniques.

Affinity screening using competitive binding with fluorine-19 hyperpolarized ligands

Kim, Y. and C. Hilty, Affinity screening using competitive binding with fluorine-19 hyperpolarized ligands. Angew Chem Int Ed Engl, 2015. 54(16): p. 4941-4.

http://www.ncbi.nlm.nih.gov/pubmed/25703090

Fluorine-19 NMR and hyperpolarization form a powerful combination for drug screening. Under a competitive equilibrium with a selected fluorinated reporter ligand, the dissociation constant (K(D)) of other ligands of interest is measurable using a single-scan Carr-Purcell-Meiboom-Gill (CPMG) experiment, without the need for a titration. This method is demonstrated by characterizing the binding of three ligands with different affinities for the serine protease trypsin. Monte Carlo simulations show that the highest accuracy is obtained when about one-half of the bound reporter ligand is displaced in the binding competition. Such conditions can be achieved over a wide range of affinities, allowing for rapid screening of non-fluorinated compounds when a single fluorinated ligand for the binding pocket of interest is known.

Dynamic nuclear polarization enhanced NMR spectroscopy for pharmaceutical formulations

Rossini, A.J., et al., Dynamic nuclear polarization enhanced NMR spectroscopy for pharmaceutical formulations. J Am Chem Soc, 2014. 136(6): p. 2324-34.

http://www.ncbi.nlm.nih.gov/pubmed/24410528

Dynamic nuclear polarization (DNP) enhanced solid-state NMR spectroscopy at 9.4 T is demonstrated for the detailed atomic-level characterization of commercial pharmaceutical formulations. To enable DNP experiments without major modifications of the formulations, the gently ground tablets are impregnated with solutions of biradical polarizing agents. The organic liquid used for impregnation (here 1,1,2,2-tetrachloroethane) is chosen so that the active pharmaceutical ingredient (API) is minimally perturbed. DNP enhancements (epsilon) of between 40 and 90 at 105 K were obtained for the microparticulate API within four different commercial formulations of the over-the-counter antihistamine drug cetirizine dihydrochloride. The different formulations contain between 4.8 and 8.7 wt % API. DNP enables the rapid acquisition with natural isotopic abundances of one- and two-dimensional (13)C and (15)N solid-state NMR spectra of the formulations while preserving the microstructure of the API particles. Here this allowed immediate identification of the amorphous form of the API in the tablet. API-excipient interactions were observed in high-sensitivity (1)H-(15)N correlation spectra, revealing direct contacts between povidone and the API. The API domain sizes within the formulations were determined by measuring the variation of epsilon as a function of the polarization time and numerically modeling nuclear spin diffusion. Here we measure an API particle radius of 0.3 mum with a single particle model, while modeling with a Weibull distribution of particle sizes suggests most particles possess radii of around 0.07 mum.

DNP Spectroscopy to characterize small molecules

The last year has seen a tremendous amount of scientific articles published in the area of Dynamic Nuclear Polarization (DNP) and hyperpolarization in general. There are several articles that demonstrate the large potential of DNP-NMR spectroscopy and I will select a few of them in the coming weeks in a combined post.

Check out these two articles that were published last year (if you haven’t done so already):

Lee, D., et al., Enhanced solid-state NMR correlation spectroscopy of quadrupolar nuclei using dynamic nuclear polarization. J Am Chem Soc, 2012. 134(45): p. 18491-4.

http://dx.doi.org/10.1021/ja307755t

Rossini, A.J., et al., Dynamic nuclear polarization NMR spectroscopy of microcrystalline solids. J Am Chem Soc, 2012. 134(40): p. 16899-908.

http://dx.doi.org/10.1021/ja308135r

Both demonstrate the large potential of DNP-NMR spectroscopy to study small molecules with only natural abundance level 13C labels. In both articles 2D 13C-13C correlation spectra of the unlabeled material are shown that would require days of signal averaging without the aid of DNP.

DNP Spectroscopy to characterize small molecules

The last year has seen a tremendous amount of scientific articles published in the area of Dynamic Nuclear Polarization (DNP) and hyperpolarization in general. There are several articles that demonstrate the large potential of DNP-NMR spectroscopy and I will select a few of them in the coming weeks in a combined post.

Check out these two articles that were published last year (if you haven’t done so already):

Lee, D., et al., Enhanced solid-state NMR correlation spectroscopy of quadrupolar nuclei using dynamic nuclear polarization. J Am Chem Soc, 2012. 134(45): p. 18491-4.

http://dx.doi.org/10.1021/ja307755t

Rossini, A.J., et al., Dynamic nuclear polarization NMR spectroscopy of microcrystalline solids. J Am Chem Soc, 2012. 134(40): p. 16899-908.

http://dx.doi.org/10.1021/ja308135r

Both demonstrate the large potential of DNP-NMR spectroscopy to study small molecules with only natural abundance level 13C labels. In both articles 2D 13C-13C correlation spectra of the unlabeled material are shown that would require days of signal averaging without the aid of DNP.

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.

http://pubs.acs.org/doi/abs/10.1021/ja308135r

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.

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.

http://pubs.acs.org/doi/abs/10.1021/ja308135r

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.

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