Fluorinated Carbohydrates as Lectin Ligands: Dissecting Glycan–Cyanovirin Interactions by Using 19F NMR Spectroscopy

NMR spectroscopy and isothermal titration calorimetry (ITC) are powerful methods to investigate ligand–protein interactions. Here, we present a versatile and sensitive fluorine NMR spectroscopic approach that exploits the ¹⁹F nucleus of ¹⁹F‐labeled carbohydrates as a sensor to study glycan binding to lectins. Our approach is illustrated with the 11 kDa Cyanovirin‐N, a mannose binding anti‐HIV lectin. Two fluoro‐deoxy sugar derivatives, methyl 2‐deoxy‐2‐fluoro‐α‐D ‐mannopyranosyl‐(1→2)‐α‐D ‐mannopyranoside and methyl 2‐deoxy‐2‐fluoro‐α‐D ‐mannopyranosyl‐(1→2)‐α‐D ‐mannopyranosyl‐(1→2)‐α‐D ‐mannopyranoside were utilized. Binding was studied by ¹⁹F NMR spectroscopy of the ligand and ¹H–¹⁵N HSQC NMR spectroscopy of the protein. The NMR data agree well with those obtained from the equivalent reciprocal and direct ITC titrations. Our study shows that the strategic design of fluorinated ligands and fluorine NMR spectroscopy for ligand screening holds great promise for easy and fast identification of glycan binding, as well as for their use in reporting structural and/or electronic perturbations that ensue upon interaction with a cognate lectin.     

Acetone‐induced polymerisation of 3‐aminopropyltrimethoxysilane (APTMS) as revealed by NMR spectroscopy

We followed the reactivity of acetone with 3‐aminopropyltrimethoxysilane, a potential organosilane coupling agent, by ¹H, ¹³C and ²⁹Si NMR spectroscopy. Selective 1D and 2D‐edited NMR experiments significantly contributed to simplify the spectral complexity of reaction solution and elucidated molecular structures within progressive reaction phases. The course of the 3‐aminopropyltrimethoxysilane reaction with acetone was shown by a progressive decrease of both reactants, and a concomitant appearance of water and methanol, due to formation of imine and hydrolysis of alkoxysilane groups, respectively. The occurrence of multiple siloxane linkages in a progressively larger cross‐linked macromolecular structure was revealed by DOSY‐NMR experiments and new signals in ²⁹Si‐NMR spectra at different reaction times. The NMR approach described here may be applied to investigate the reactivity of other γ‐aminopropylalkoxysilanes and contribute to define procedures for the preparation of silica‐based materials. Copyright © 2014 John Wiley & Sons, Ltd.     

Chemical‐Shift‐Resolved 19F NMR Spectroscopy between 13.5 and 135 MHz: Overhauser–DNP‐Enhanced Diagonal Suppressed Correlation Spectroscopy

Overhauser–DNP‐enhanced homonuclear 2D ¹⁹F correlation spectroscopy with diagonal suppression is presented for small molecules in the solution state at moderate fields. Multi‐frequency, multi‐radical studies demonstrate that these relatively low‐field experiments may be operated with sensitivity rivalling that of standard 200–1000 MHz NMR spectroscopy. Structural information is accessible without a sensitivity penalty, and diagonal suppressed 2D NMR correlations emerge despite the general lack of multiplet resolution in the 1D ODNP spectra. This powerful general approach avoids the rather stiff excitation, detection, and other special requirements of high‐field ¹⁹F NMR spectroscopy.     

Single‐Scan Multidimensional NMR Analysis of Mixtures at Sub‐Millimolar Concentrations by using SABRE Hyperpolarization

Signal amplification by reversible exchange (SABRE) is a promising method to increase the sensitivity of nuclear magnetic resonance (NMR) experiments. However, SABRE‐enhanced ¹H NMR signals are short lived, and SABRE is often used to record 1D NMR spectra only. When the sample of interest is a complex mixture, this results in severe overlaps for ¹H spectra. In addition, the use of a co‐substrate, whose signals may obscure the ¹H spectra, is currently the most efficient way to lower the detection limit of SABRE experiments. Here, we describe an approach to obtain clean, SABRE‐hyperpolarized 2D ¹H NMR spectra of mixtures of small molecules at sub‐millimolar concentrations in a single scan. The method relies on the use of para‐hydrogen together with a deuterated co‐substrate for hyperpolarization and ultrafast 2D NMR for acquisition. It is applicable to all substrates that can be polarized with SABRE.     

Spectral study of a new dihydroisocoumarin

A novel dihydroisocoumarin, 3,4‐dihydro‐6,8‐dihydroxy‐3‐(2′‐acetyl‐3′‐hydroxy‐5′‐methoxy)methyl‐1H‐[2]benzopyran‐1‐one, was isolated from the chloroform extract of the sap of the traditional herb Aloe vera. Its structure was determined by high‐resolution negative fast atom bombardment mass spectrometry (MS), 2D NMR spectroscopy and x‐ray crystallography. The molecular structure was elucidated by 2D NMR analysis. The complete assignment of the ¹H and ¹³C NMR spectra of this compound was performed by using ¹H detected one‐bond heteronuclear multiple quantum correlation (HMQC) and long‐range (two and three bonds) heteronuclear multiple quantum bond correlation (HMBC) experiments. Detailed analyses of the one‐ and two‐dimensional NMR techniques are presented in additional to the spectral properties (MS, IR and UV). Copyright © 2003 John Wiley & Sons, Ltd.     

Technical and practical aspects of 19F NMR‐based screening: toward sensitive high‐throughput screening with rapid deconvolution

The technical and practical aspects of ¹⁹F NMR‐based screening against a macromolecular target are analyzed in detail. A novel method utilizing the relaxation of ¹⁹F homonuclear double quantum coherence is proposed for performing NMR‐based binding assays in a direct‐ or competition‐mode format. A combined strategy based on ¹⁹F NMR chemical shift prediction, 2D ¹⁹F NMR DOSY, and 2D ¹⁹F–¹H NMR long‐range COSY experiments is presented for the deconvolution of complex mixtures of fluorinated molecules generated by either addition of single compounds or by chemical synthesis. The approaches presented here allow the screening of complex mixtures, even in the case where the exact composition is not known, and the rapid identification of the binders contained in the mixtures. Copyright © 2012 John Wiley & Sons, Ltd.     

Total assignment of 1H and 13C NMR spectra of three triterpene saponins from roots of Silene vulgaris (Moench) Garcke

The assignments of ¹H and ¹³C NMR spectra for three new triterpene saponins from Silene vulgaris (gypsogenin 3‐O‐glucuronide, quillaic acid 3‐O‐glucuronide, and gypsogenin 3‐O‐glycoside) are reported. In addition to 1D NMR methods, 2D NMR techniques (COSY, HSQC, HMBC, and HSQC‐TOCSY) were used for the assignments. Copyright © 2002 John Wiley & Sons, Ltd.     

The Low Polar Constituents from Bidens Pilosa L. var. Minor (Blume) Sherff

Anew and 20 known compounds were isolated from Bidens pilosa L. var. minor (Blume) Sherff. The new compound was determined as 7‐phenyl‐hepta‐4,6‐diyne‐2‐ol by various physical techniques (MS, IR, UV, ¹H‐, ¹³C‐NMR, and 2D‐NMR).     

A new coumarin from Perezia hebeclada

Methanol extracts from Perezia hebeclada roots yielded the new 8‐β‐D ‐glucopyranosyloxy‐4‐methoxy‐5‐methylcoumarin ( 1 ) together with the known 4‐β‐D ‐glucopyranosyloxy‐5‐methylcoumarin ( 2 ). Their structures were determined and verified, respectively, by MS and NMR studies, including 1D and 2D experiments. Two ¹³C NMR signals of the sugar residue of 2 were reassigned. Copyright © 2003 John Wiley & Sons, Ltd.     

Structural analyses of mannose pentasaccharide of high mannose type oligosaccharides by 1D and 2D NMR spectroscopy

NMR spectroscopy is a very important and useful method for the structural analysis of oligosaccharides, despite its low sensitivity. We first applied conventional measuring methods, 2D DQF COSY, ¹H–¹³C HSQC, and ¹H–¹³C HMBC, and also the Double Pulsed Field Gradient Spin Echo (DPFGSE)‐TOCSY and DPFGSE‐NOESY/ROESY techniques to analyze a branched mannose pentasaccharide as a model of high mannose type N‐glycans in natural abundance. The NMR spectra of the model compound are very complex and difficult to analyze owing to overlapping signals. The superior selective irradiation capability of the DPFGSE technique is useful for fine structural and conformational analyses of such complex oligosaccharides. We here introduce a novel technique called DPFGSE‐Double‐Selective Population Transfer (SPT)‐Difference and DPFGSE‐NOE/ROE‐SPT‐Difference spectroscopy. The DPFGSE‐Double‐SPT‐Difference method involves irradiation of two peaks from one proton and the subtraction of higher and lower peaks from each spectrum. The DPFGSE‐NOE/ROE‐SPT‐Difference method involves the transfer of the magnetization polarized by NOE/ROE from the nuclei to the spin‐coupled nuclei through scalar spin–spin interaction using the SPT method. Even if the signals in the NMR spectra overlap, each signal can be accurately assigned. In particular, DPFGSE‐NOE/ROE‐SPT‐Difference is very useful for identifying sugar connectivity. Copyright © 2012 John Wiley & Sons, Ltd.     

Kinetics of Core‐Shell Nanoparticle Formation by Two‐Dimensional Nuclear Magnetic Resonance

We have studied the kinetics of polymeric nanoparticle formation for poly(styrene‐block‐4‐vinylpyridine) [P(S‐b‐4‐VPy)], chains in a non‐selective solvent using 1,4‐dibromobutane (DBB) as a cross‐linker by means of different nuclear magnetic resonance (NMR) spectroscopy techniques. The kinetic process was followed using ¹H, ¹³C, and 2‐D Heteronuclear Single Quantum Correlation (HSQC) NMR experiments. The kinetic data obtained from 2‐D HSQC and ¹H NMR experiments were in good agreement between them, proving the reliability of the 2‐D HSQC NMR technique for the in situ study of the kinetics of core‐shell nanoparticle formation. A value of 1.5 × 10⁻⁵ s⁻¹ was determined for the apparent kinetic constant of the P(S‐b‐4‐VPy)‐DBB core‐shell nanoparticle formation process.

     

Complete assignments of the 1H and 13C NMR spectra of five rearranged abietane diterpenoids

An NMR study of five highly functionalized and rearranged abietane diterpenoids is described. In addition to 1D NMR methods, including 1D NOESY spectra, 2D shift‐correlated experiments [¹H, ¹³C‐gHSQC‐¹J (C,H) and ¹H, ¹³C‐gHMBC‐ⁿJ (C,H) (n = 2 and 3)] were used for the complete and unambiguous ¹H and ¹³C chemical shift assignments of these substances. Copyright © 2002 John Wiley & Sons, Ltd.     

Synthesis,crystal structures,spectral FT‐IR,NMR and UV–Vis investigations and Hirshfeld surface analysis of two new 2‐hydroxyethyl‐substituted N‐heterocyclic carbene precursors

This study discusses the synthesis of two new 2‐hydroxyethyl substituted N‐heterocyclic carbene (NHC) precursors. The NHC precursors were prepared from 1‐(alkyl/aryl)benzimidazole and alkyl halides. They were characterized using ¹H NMR, ¹³C NMR, FT‐IR, UV–Vis spectroscopy, and elemental analysis techniques. Molecular and crystal structures of 1 and 2 were determined using the single‐crystal X‐ray diffraction method. Crystal structure of the compounds features NHC precursors and chloride anions. Additionally in 2 , the asymmetric unit has a water molecule, which forms a tetrameric chloride‐hydrate assembly with the chloride anion. The chloride anions play an important role in the stabilization of crystal structures to form a two‐dimensional supramolecular architecture. The 3D Hirshfeld surface and the associated 2D fingerprint plots were also drawn to gain insights into the behavior of the interactions in the compounds.     

An Approach to the Synthesis of Spiro[indene‐pyridoisoquinoline] Derivatives via 1,4‐Dipolar Cycloaddition of Isoquinoline and Acetylene Esters,and (1,3‐Dihydro‐1,3‐dioxo‐2H‐inden‐2‐ylidene)malononitrile

A concise and efficient approach to the spiro‐tetrahydroisoquinoline derivatives has been developed by 1,4‐dipolar cycloaddition of zwitterions resulting from isoquinoline and acetylene esters and (1,3‐dihydro‐1,3‐dioxo‐2H‐inden‐2‐ylidene)malononitrile in MeCN at room temperature. The significance of this method lies in good yields and ease of product purification, and no inert atmosphere is required. The structures of the products were confirmed spectroscopically (IR, ¹H‐ and ¹³C‐NMR, and EI‐MS) and by elemental analyses. A plausible mechanism for this reaction is proposed (Scheme).     

Structure elucidation and antibacterial activity of a new coumarinolignoid from Daphne gnidium L.

Phytochemical investigation of the dichloromethane extract from the leaves of Daphne gnidium L. afforded 2‐(4‐hydroxy‐3,5‐dimethoxyphenyl)‐3‐hydroxymethyl‐2,3‐dihydro‐1,4,5‐trioxaphenanthren‐6‐one, and its structure was determined by a combination of spectral methods (1D and 2D ¹H, ¹³C NMR and HRMALDI and EI mass spectrometry). The new compound was found to possess moderate antibacterial activity against Staphylococcus aureus and Bacillus cereus. However, it showed no cytotoxic activity against KB cells. Copyright © 2002 John Wiley & Sons, Ltd.     

Advanced solvent signal suppression for the acquisition of 1D and 2D NMR spectra of Scotch Whisky

A simple and robust solvent suppression technique that enables acquisition of high‐quality 1D ¹H nuclear magnetic resonance (NMR) spectra of alcoholic beverages on cryoprobe instruments was developed and applied to acquire NMR spectra of Scotch Whisky. The method uses 3 channels to suppress signals of water and ethanol, including those of ¹³C satellites of ethanol. It is executed in automation allowing high throughput investigations of alcoholic beverages. On the basis of the well‐established 1D nuclear Overhauser spectroscopy (NOESY) solvent suppression technique, this method suppresses the solvent at the beginning of the pulse sequence, producing pure phase signals minimally affected by the relaxation. The developed solvent suppression procedure was integrated into several homocorrelated and heterocorrelated 2D NMR experiments, including 2D correlation spectroscopy (COSY), 2D total correlation spectroscopy (TOCSY), 2D band‐selective TOCSY, 2D J‐resolved spectroscopy, 2D ¹H, ¹³C heteronuclear single‐quantum correlation spectroscopy (HSQC), 2D ¹H, ¹³C HSQC‐TOCSY, and 2D ¹H, ¹³C heteronuclear multiple‐bond correlation spectroscopy (HMBC). A 1D chemical‐shift‐selective TOCSY experiments was also modified. The wealth of information obtained by these experiments will assist in NMR structure elucidation of Scotch Whisky congeners and generally the composition of alcoholic beverages at the molecular level.     

2－脱氧－烟酰胺基－β－D－氨基葡萄糖的合成和表征

A new nicotinic acid derivative,2-deoxy-2-nicotinoylamido-β-D-glucopyranose, was synthesized with β-configuration exclusively. The structure and properties of the product were characterized by ^1H NMR, PT-IR, MS, DSC and polarimeter. The details of ^1H NMR spectrum and the mass spectrum proved that there are a great amount of hydrogen bonds in the product.     

Positional Enrichment by Proton Analysis (PEPA): A One‐Dimensional 1H‐NMR Approach for 13C Stable Isotope Tracer Studies in Metabolomics

A novel metabolomics approach for NMR‐based stable isotope tracer studies called PEPA is presented, and its performance validated using human cancer cells. PEPA detects the position of carbon label in isotopically enriched metabolites and quantifies fractional enrichment by indirect determination of ¹³C‐satellite peaks using 1D‐¹H‐NMR spectra. In comparison with ¹³C‐NMR, TOCSY and HSQC, PEPA improves sensitivity, accelerates the elucidation of ¹³C positions in labeled metabolites and the quantification of the percentage of stable isotope enrichment. Altogether, PEPA provides a novel framework for extending the high‐throughput of ¹H‐NMR metabolic profiling to stable isotope tracing in metabolomics, facilitating and complementing the information derived from 2D‐NMR experiments and expanding the range of isotopically enriched metabolites detected in cellular extracts.     

Stacking Structure of Confined 1‐Butanol in SBA‐15 Investigated by Solid‐State NMR Spectroscopy

Understanding the complex thermodynamic behavior of confined amphiphilic molecules in biological or mesoporous hosts requires detailed knowledge of the stacking structures. Here, we present detailed solid‐state NMR spectroscopic investigations on 1‐butanol molecules confined in the hydrophilic mesoporous SBA‐15 host. A range of NMR spectroscopic measurements comprising of ¹H spin–lattice (T₁), spin–spin (T₂) relaxation, ¹³C cross‐polarization (CP), and ¹H,¹H two‐dimensional nuclear Overhauser enhancement spectroscopy (¹H,¹H 2D NOESY) with the magic angle spinning (MAS) technique as well as static wide‐line ²H NMR spectra have been used to investigate the dynamics and to observe the stacking structure of confined 1‐butanol in SBA‐15. The results suggest that not only the molecular reorientation but also the exchange motions of confined molecules of 1‐butanol are extremely restricted in the confined space of the SBA‐15 pores. The dynamics of the confined molecules of 1‐butanol imply that the ¹H,¹H 2D NOESY should be an appropriate technique to observe the stacking structure of confined amphiphilc molecules. This study is the first to observe that a significant part of confined 1‐butanol molecules are orientated as tilted bilayered structures on the surface of the host SBA‐15 pores in a time‐average state by solid‐state NMR spectroscopy with the ¹H,¹H 2D NOESY technique.