Low-power composite CPMG HSQMBC experiment for accurate measurement of long-range heteronuclear coupling constants

A modified version of CPMG-HSQMBC pulse scheme is presented for the measurement of long-range heteronuclear coupling constants. The method implements adiabatic inversion and refocusing pulses on the heteronucleus. Low-power composite 180° XY-16 CPMG pulse train is applied on both proton and X nuclei during the evolution of long-range couplings to eliminate phase distortions due to co-evolution of homonuclear proton-proton couplings. The pulse sequence yields pure absorption antiphase multiplets allowing precise and direct measurement of the (n)J(XH) coupling constants regardless from the size of the proton-proton couplings. The applicability of the method is demonstrated using strychnine as a model compound. The selective 1D version of the method is also presented.     

Measurement of long‐range proton–carbon coupling constants from pure in‐phase 1D multiplets

The SELective INverse detection of carbon–proton CORrelation pulse sequence that yields a 1D spectrum of a proton directly bonded to a selected carbon resonance has been converted into a proton and carbon double‐selective variant that provides a ¹H spectrum of a selected proton that is long‐range coupled to a specific carbon resonance. The resulting 1D proton multiplet exhibits a pure absorptive in‐phase lineshape for precise measurement of specific long‐range proton–carbon coupling constants in small organic molecules at natural abundance. Copyright © 2011 John Wiley & Sons, Ltd.     

Simultaneous measurement of N?H and Cα?Hα coupling constants in proteins

We present a pulse sequence for the simultaneous measurement of N? H and Cα? Hα couplings in double‐labeled proteins from 2D spectra. The proposed sequence, a modification of the HN(CO)CA experiment, combines the J‐modulation method and the IPAP scheme. The couplings can be readily retrieved from a series of 2D ¹⁵N? ¹H correlation spectra, differing in the time point at which a ¹H 180° pulse is applied. This induces an intensity modulation of the ¹⁵N? ¹H correlation peaks with the Cα? Hα coupling. The Cα? Hα coupling is then obtained by fitting the observed intensities to the modulation equation. The N? H coupling is measured in each member of the set from peak‐to‐peak separations in the IPAP subspectra. The pulse sequence is experimentally verified with a sample of ¹⁵N/¹³C‐enriched ubiquitin. Copyright © 2009 John Wiley & Sons, Ltd.     

1D NMR Homodecoupled 1H Spectra with Scalar Coupling Constants from 2D NemoZS‐DIAG Experiments

A two‐dimensional liquid‐state NMR experiment cleanly separating chemical shifts and scalar couplings information is introduced. This DIAG experiment takes advantage of a drastic reduction of the spectral window in the indirect dimension to be quickly recorded and of a new non‐equidistant modulation of the selective pulse to improve the sensitivity of the broadband homodecoupling Zangger–Sterk sequence element by one order of magnitude. A simple automatic analysis results in 1D spectra displaying singlets and lists of the scalar couplings for first‐order multiplets. This facilitates the analysis of 1D spectra by resolving multiplets based on their differences in chemical shifts and coupling structures.     

New routes to the detection of relaxation allowed coherence transfer in paramagnetic molecules

A new pulse sequence is proposed to measure cross correlation rates between 1H Curie spin relaxation and 1H-15N or 1H-13C dipole-dipole coupling (%@mt;sys@%Gamma%@sx@%H,HX%@be@%CS,DD%@sxx@%%@mx@% ) in paramagnetic systems. The new sequence has been conceived to obtain quantitative measurements of cross correlation rates in the close proximity to the paramagnetic center, preventing the loss of information due to fast transverse relaxation. The approach was tested on the dicalcium protein calbindin D9k in which Ca2+ has been substituted at site II with Ce3+. At variance with previously reported experiments, all peaks observed in HSQC experiments tailored to paramagnetic signals give quantitative estimates of %@mt;sys@%Gamma%@sx@%H%@ital@%,%@rsf@%HX%@be@%CS%@ital@%,%@rsf@%DD%@sxx@%%@mx@% . This is crucial to refine the immediate proximity of the metal ion.     

Fluorescence-detected two-dimensional electronic coherence spectroscopy by acousto-optic phase modulation

Two-dimensional electronic coherence spectroscopy (ECS) is an important method to study the coupling between distinct optical modes of a material system. Such studies often involve excitation using a sequence of phased ultrashort laser pulses. In conventional approaches, the delays between pulse temporal envelopes must be precisely monitored or maintained. Here, we introduce a new experimental scheme for phase-selective nonlinear ECS, which combines acousto-optic phase modulation with ultrashort laser excitation to produce intensity modulated nonlinear fluorescence signals. We isolate specific nonlinear signal contributions by synchronous detection, with respect to appropriately constructed references. Our method effectively decouples the relative temporal phases from the pulse envelopes of a collinear train of four sequential pulses. We thus achieve a robust and high signal-to-noise scheme for phase-selective ECS to investigate the resonant nonlinear optical response of photoluminescent systems. We demonstrate the validity of our method using a model quantum three-level system-atomic Rb vapor. Moreover, we show how our measurements determine the resonant complex-valued third-order susceptibility.     

High-resolution NMR spectra under inhomogeneous fields via intermolecular double-quantum coherences

High-resolution NMR spectroscopy is a powerful tool for analyzing molecular structures and compositions. Line-widths of conventional liquid NMR signals are directly proportional to the overall magnetic field inhomogeneity the sample experiences. In many circumstances, spatial and temporal homogeneity of the magnetic field is degraded. In this paper, a modified pulse sequence based on intermolecular double-quantum coherences (iDQCs) was proposed to obtain 1D high-resolution NMR spectra under inhomogeneous fields using 2D acquisition. Analytical expressions were derived from the intermolecular multiple-quantum coherence (iMQC) treatments. Both experimental and simulated spectra provide high-resolution 1D projection spectra similar to conventional 1D high-resolution spectra. Moreover, the apparent J coupling constants are threefold magnified, which allows a more accurate measurement of small J coupling constants.     

Selective measurement of heteronuclear 1H-13C dipolar couplings in motionally heterogeneous semicrystalline polymer systems

A pulse sequence for the selective recoupling of heteronuclear dipolar interactions in mobile amorphous phase of powdered semicrystalline polymers is described. 1H-13C dipolar interactions are selectively measured by PISEMA-type sequence. Selection of 13C magnetization originating from amorphous phase is achieved by a train of saturation pulses followed by a short delay and a direct excitation pulse on 13C spins. The development of undesired net 13C magnetization during the recoupling sequence is prevented by the efficient "reverse" 13C --> 1H cross-polarization. The efficacy of the 2D method to measure 1H-13C dipolar couplings selectively for mobile components is demonstrated on powdered crystalline L-alanine, semicrystalline polyethylene, and nanocomposite polyamide-6/montmorillonite.     

Frequency-swept HMQC sequences for high-throughput NMR analysis

We describe here new versions of the DEPT phase-encoded HMQC experiment that offer robust performance and improved sensitivity. The new sequences rely on frequency-swept proton and carbon pulses to minimize signal losses from miscalibrated pulses while providing 'J compensation' to optimize the signal strength over a range of heteronuclear coupling constants. By including both proton and carbon-swept pulses, the new sequences also offer an additional signal gain of roughly 10% over well-calibrated hard-pulse experiments. The new sequences also demonstrate that one can construct a sequence that incorporates both 90 degrees and 180 degrees frequency-swept pulses. Although individual pulses in the sequence cause severe phase roll, the phase roll can be eliminated by the proper choice of pulse lengths and sweep directions.     

盐酸L-肉碱和盐酸乙酰L-肉碱的合成

L-肉碱(维生素BT)和乙酰L-肉碱是长链脂肪酸运输的载体, 广泛用于医药、保健食品及饲料等行业. 本工作介绍了一种化学不对称合成盐酸L-肉碱和盐酸乙酰L-肉碱的方法. 以自制的手性噁唑硼烷为催化剂, 将起始原料γ-氯乙酰乙酸乙酯不对称催化氢化还原成(R)-γ-氯-β-羟基丁酸乙酯, 这一步的化学收率和光学收率都较高. 然后将(R)-γ-氯-β-羟基丁酸乙酯转化为盐酸L-肉碱和盐酸乙酰L-肉碱. 合成盐酸L-肉碱只需两步, 总收率为68.3%, ee值可达94.6%; 合成盐酸乙酰L-肉碱需三步, 总收率为62.6%, ee值可达96.8%. 还考察了反应温度、催化剂及还原剂的量等因素对γ-氯乙酰乙酸乙酯的不对称氢化还原的化学产率和光学收率的影响.     

Heteronuclear two-bond correlation: suppressing heteronuclear three-bond or higher NMR correlations while enhancing two-bond correlations even for vanishing 2J(CH)

A novel two-dimensional NMR pulse sequence, H2BC, for long-range correlation of 1H and 13C nuclei is presented. The experiment has several attractive features compared to the widely used HMBC experiment, for example, (a) typically strong enhancement of correlations over two bonds while suppressing those over more bonds, that is, resolving ambiguities in standard HMBC spectra and showing two-bond correlations not present in HMBC spectra, (b) independence of long-range 1H-13C coupling constants, (c) full homo- and heteronuclear decoupling in the indirect dimension and heteronuclear decoupling in the acquisition dimension, (d) pure 2D absorption peak shapes, and (e) a pulse sequence duration significantly shorter than that of HMBC. The experiment is quite complementary to HMBC and does not effect correlations to quaternary carbons that must be obtained by HMBC.     

一种新的测定寡糖链中糖体亚谱的NMR方法

报道一种新的测定寡糖链糖体亚谱的NMR方法, 称为MMDY混合脉冲梯度选择一维HOHAHA法. 本方法采用MMDY混合脉冲与双梯度自旋回波脉冲相结合, 为解决高度重叠的¹H NMR谱图分析困难提供了有力手段. 本方法的主要特点是: 采用双梯度自旋回波脉冲进行选择激发, 与差谱为基础的选择激发方法相比不但可节省测定时间, 而且谱图干净, 没有差谱伪峰; MMDY混合脉冲的磁化接力相干转移非常有效, 远程磁化转移信号的灵敏度高, 对于偶合常数大约为1 Hz的微弱偶合自旋体系, 如阿拉伯糖和鼠李糖, 也能够得到完整的亚谱; 梯度脉冲只是用于相干路径选择, 灵敏度比较高, 并且纯吸收形的高质量亚谱不经过复杂的Z-滤波步骤就可以快速得到, 因此比较适用于常规样品分析. 以苷类化合物为例讨论本方法的特点以及在谱峰高度的寡糖结构测定中的应用.     

(119/117/115)Sn low-abundance single-transition correlation spectroscopy (LASSY): sensitivity-enhanced homonuclear correlation experiments for Sn NMR

We report the implementation of our novel rare-spin homonuclear correlation experiment, namely, Low-Abundance Single-transition correlation SpectroscopY (LASSY), for (119/117/115)Sn NMR at natural abundance. Our pulse sequence results in diagonal suppressed COSY-style display and outperforms the optimal homonuclear correlation experiment for rare spins, which involves double quantum evolution (INADEQUATE CR). The new experiment maximizes efficiency both in respect of pulse transformations as well as relaxation effects, and gives rise to a simplified two-dimensional (2D) spectrum with considerably reduced crowding, exhibiting only one transition in each cross peak, instead of four. Performance optimization of LASSY is carried out in light of the relatively 'large' line widths typical of Sn NMR in solution state. The superior performance of the sequence is demonstrated on dimeric tetraorganodistannoxane samples.     

New DEFT sequences for the acquisition of one-dimensional carbon NMR spectra of small unlabelled molecules

The acquisition time and quality of 1D 13C{1H} spectra can be improved substantially by using a modified driven equilibrium Fourier transform (DEFT) sequence, which is specifically designed to compensate for the effects of B1 inhomogeneity, pulse miscalibration and frequency offsets. The new sequence, called uniform driven equilibrium Fourier transform (UDEFT), returns the carbon magnetization with a high accuracy along its equilibrium position after each transient is complete. Thus, the sequence allows the use of relaxation delays (RD), which are much shorter than the carbon T1 of the molecule, thereby speeding up the acquisition process of 1D 13C{1H} spectra. To achieve this level of performance, UDEFT employs a refocusing element constituted by a composite adiabatic carbon pulse surrounded by two 90 degrees carbon pulses whose phases are designed to compensate for 90 degrees pulse miscalibrations in an MLEV manner (90 degrees+x-tau(FID)-180+y(Adia)-tau-90 degrees+x-180 degrees+x(Adia)). A version of the UDEFT sequence allows recording 1D 13C{1H} spectra devoid of heteronuclear NOE by using a matched adiabatic 1H decoupling scheme where an even number of 180 degrees adiabatic pulses is applied during the UDEFT module. Spectra of a solution of 300 mM camphor that contains some carbon nuclei with very long T1 relaxation times (90 s and 78 s) were acquired with 128 scans in 10 min using a 5 s relaxation delay.     

Suppression of phase and amplitude J(HH) modulations in HSQC experiments

The amplitude and the phase of cross peaks in conventional 2D HSQC experiments are modulated by both proton–proton, J(HH), and proton–carbon, ¹J(CH), coupling constants. It is shown by spectral simulation and experimentally that J(HH) interferences are suppressed in a novel perfect‐HSQC pulse scheme that incorporates perfect‐echo INEPT periods. The improved 2D spectra afford pure in‐phase cross peaks with respect to ¹J(CH) and J(HH), irrespective of the experiment delay optimization. In addition, peak volumes are not attenuated by the influence of J(HH), rendering practical issues such as phase correction, multiplet analysis, and signal integration more appropriate. Copyright © 2014 John Wiley & Sons, Ltd.     

Fully Resolved NMR Correlation Spectroscopy

A new correlation experiment cited as “push‐G‐SERF” is reported. In the resulting phased 2D spectrum, the chemical shift information is selected along the direct dimension, whereas scalar couplings involving a selected proton nucleus are edited in the indirect domain. The robustness of this pulse sequence is demonstrated on compounds with increasing structural and spectral complexity, using state‐of‐the‐art spectrometers. It allows for full resolution of both dimensions of the spectrum, yielding a straightforward assignment and measurement of the coupling network around a given proton in the molecule. This experiment is intended for chemists who want to address efficiently the structural analysis of molecules with an overcrowded spectrum.     

Pseudomultidimensional NMR by spin-state selective off-resonance decoupling

An alternate technique for accurately monitoring the chemical shift in multidimensional NMR experiments using spin-state selective off-resonance decoupling is presented here. By applying off-resonance decoupling on spin S during acquisition of spin I, we scaled the scalar coupling J(I,S) between the spins, and the residual scalar coupling turns out to be a function of the chemical shift of spin S. Thus, the chemical shift information of spin S is indirectly retained, without an additional evolution period and the accompanying polarization transfer elements. The detection of the components of the doublet using spin-state selection enables an accurate measurement of the residual scalar coupling and a precise value for the chemical shift, concomitantly. The spin-state selection further yields two subspectra comprising either one of the two components of the doublet and thereby avoiding the overlap problems that arise from off-resonance decoupling. In general, spin-state selective off-resonance decoupling can be incorporated into any pulse sequence. Here, the concept of spin-state selective off-resonance decoupling is applied to 3D (13)C or (15)N-resolved [(1)H,(1)H]-NOESY experiments, adding the chemical shift of the heavy atom attached to the hydrogen ((13)C or (15)N nuclei) with high resolution resulting in a pseudo-4D. These pseudo-4D heavy-atom resolved [(1)H, (1)H]-NOESY experiments contain chemical shift information comparable to that of 4D (13)C or (15)N-resolved [(1)H,(1)H]-NOESY, but with an increase in chemical shift resolution by 1-2 orders of magnitude.     

Site-selective screening by NMR spectroscopy with labeled amino acid pairs

A new method for site-selective screening by NMR is presented. The core of the new method is the dual amino acid sequence specific labeling technique. Amino acid X is labeled with (13)C and amino acid Y is labeled with (15)N. Provided only one XY pair occurs in the amino acid sequence, only one signal in the 1D carbonyl (13)C spectrum will display a splitting due to the (1)J(C'N) coupling. Using this labeling strategy it is possible to screen selectively for binding to a selected epitope without the need for sequence specific assignments. An HNCO spectrum (1D or 2D) can be used either directly as a screening experiment or indirectly to identify what signals to monitor in a 2D (1)H-(15)N correlation spectrum. Chemical shift perturbations upon addition of a potential ligand are easily detected even for large proteins due to the reduced spectral complexity resulting from the use of a selectively labeled sample. The new technique is demonstrated on the human adipocyte fatty acid binding protein FABP-4. Due to the reduced spectral complexity, the method should be applicable to larger proteins than are conventional methods.     

An experimental study of decoupling sequences for multiple-quantum and high-resolution MAS experiments in solid-state NMR

Recently, a sequence for heteronuclear dipolar decoupling in solid-state NMR, namely SWf-TPPM, was introduced by us. Under magic-angle spinning (MAS), the decoupling efficiency of the sequence was unaffected over a range of values for various experimental parameters such as the pulse length, pulse phase, and 1H resonance offset. We here demonstrate its use in multiple-quantum (MQ) and high-resolution (HR) MAS experiments. This sequence further improves the MQMAS spectra compared to the earlier reported decoupling sequences with improved immunity to any missets of the pulse length, pulse phase and decoupler offset. In contrast, for HRMAS, the simple CW scheme is as efficient as any of the decoupling schemes that were studied.     

Conformations of N-acetyl-L-prolinamide by two-dimensional infrared spectroscopy

Femtosecond two-dimensional infrared (2D IR) spectroscopy has been applied to study the conformations of a model dipeptide, N-acetyl-L-prolinamide (AcProNH2) in deuterated chloroform (CDCl3). Spectral features in the amide-I and -II regions are obtained by rephasing (R), nonrephasing (NR), and reverse photon echo (RPE) pulse sequences with two polarization conditions. The 2D spectra obtained by the RPE and NR sequences with (0, 0, 0, 0) polarization reveal new spectral features associated with the multiple conformers of AcProNH2 that are difficult to discern using R sequence and linear-IR spectroscopy. The high resolving power of the RPE sequence comes from destructive interference between the positive and negative peaks of nearby vibrators, similar to the NR sequence. The RPE response functions that are useful for 2D spectral simulations are evaluated, including the effects of vibrational frequency correlations. The 2D spectra obtained with (45, -45, 90, 0) polarization exhibit clear cross-peak patterns in the off-diagonal region for the R and RPE sequences but in the diagonal region for the NR sequence. These patterns, free from strong diagonal contributions, are crucial for structure determination. DFT calculations, normal-mode analysis, Hessian matrix reconstruction, and vibrational exciton Hamiltonian diagonalization yield molecular parameters needed for quantitative simulations of 2D spectra: angles between transition dipoles, coupling constants, and off-diagonal anharmonicities of the amide-I and -II modes are obtained for solvated trans-C7 and cis structures and for gas-phase trans conformers in the region of phi = -120 degrees to 0 degrees and psi = -100 degrees to 180 degrees in the Ramachandran space. Systematic simulations based on a 4:1 population ratio of the solvated trans-C7 and cis structures reproduce well the 2D spectral features obtained at both polarization conditions. However, better agreement between the experimental and simulated cross-peak patterns can be reached if the dihedral angles of the major trans conformer are close to (phi, psi) = (-80 degrees , 100 degrees ). Our results suggest that the major conformer of AcProNH2 in CDCl3 deviates from the gas-phase global minimum, the trans-C7 form, to an extended intermediate between the C7 and polyproline-II structure. These results are discussed in relationship with earlier findings obtained by NMR, transient IR studies, and MD simulations.