Spatially Localized Two-Dimensional J-Resolved NMR Spectroscopy via Intermolecular Double-Quantum Coherences for Biological Samples at 7 T

Chunhua Tan; Shuhui Cai; Yuqing Huang

doi:10.1371/journal.pone.0134109

Abstract

Background and Purpose

Magnetic resonance spectroscopy (MRS) constitutes a mainstream technique for characterizing biological samples. Benefiting from the separation of chemical shifts and J couplings, spatially localized two-dimensional (2D) J-resolved spectroscopy (JPRESS) shows better identification of complex metabolite resonances than one-dimensional MRS does and facilitates the extraction of J coupling information. However, due to variations of macroscopic magnetic susceptibility in biological samples, conventional JPRESS spectra generally suffer from the influence of field inhomogeneity. In this paper, we investigated the implementation of the localized 2D J-resolved spectroscopy based on intermolecular double-quantum coherences (iDQCs) on a 7 T MRI scanner.

Materials and Methods

A γ-aminobutyric acid (GABA) aqueous solution, an intact pig brain tissue, and a whole fish (Harpadon nehereus) were explored by using the localized iDQC J-resolved spectroscopy (iDQCJRES) method, and the results were compared to those obtained by using the conventional 2D JPRESS method.

Results

Inhomogeneous line broadening, caused by the variations of macroscopic magnetic susceptibility in the detected biological samples (the intact pig brain tissue and the whole fish), degrades the quality of 2D JPRESS spectra, particularly when a large voxel is selected and some strongly structured components are included (such as the fish spinal cord). By contrast, high-resolution 2D J-resolved information satisfactory for metabolite analyses can be obtained from localized 2D iDQCJRES spectra without voxel size limitation and field shimming. From the contrastive experiments, it is obvious that the spectral information observed in the localized iDQCJRES spectra acquired from large voxels without field shimming procedure (i.e. in inhomogeneous fields) is similar to that provided by the JPRESS spectra acquired from small voxels after field shimming procedure (i.e. in relatively homogeneous fields).

Conclusion

The localized iDQCJRES method holds advantage for recovering high-resolution 2D J-resolved information from inhomogeneous fields caused by external non-ideal field condition or internal macroscopic magnetic susceptibility variations in biological samples, and it is free of voxel size limitation and time-consuming field shimming procedure. This method presents a complementary way to the conventional JPRESS method for MRS measurements on MRI systems equipped with broad inner bores, and may provide a promising tool for in vivo MRS applications.

Citation: Tan C, Cai S, Huang Y (2015) Spatially Localized Two-Dimensional J-Resolved NMR Spectroscopy via Intermolecular Double-Quantum Coherences for Biological Samples at 7 T. PLoS ONE 10(7): e0134109. https://doi.org/10.1371/journal.pone.0134109

Editor: Mehdi Mobli, The University of Queensland, AUSTRALIA

Received: December 23, 2014; Accepted: July 6, 2015; Published: July 24, 2015

Copyright: © 2015 Tan et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited

Data Availability: All relevant data are within the paper.

Funding: This work was supported by the National Natural Science Foundation of China under Grants 11205129 (YH) and 11275161 (SC). The URL for the grants is http://www.nsfc.gov.cn/. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Competing interests: The authors have declared that no competing interests exist.

Introduction

Magnetic resonance spectroscopy (MRS) is a powerful tool for investigating chemical compositions and elucidating molecular structures. It enables us to reveal valuable molecule-level information, such as chemical shifts, J couplings, and multiplet patterns. The spectral information, complementary to the morphological information provided by magnetic resonance imaging (MRI), is useful for analyses of biological samples [1, 2]. Due to its efficacy, MRS shows wide applications in a variety of fields [3–6]. The localized one-dimensional (1D) point-resolved spectroscopy (PRESS) is a common MRS method for practical applications with the advantage of fast acquisition [7, 8]. However, spectral congestion is generally observed in 1D PRESS spectra of biological samples because numerous resonances from various metabolites are involved in a single spectral dimension. In addition, the intrinsic magnetic susceptibility variations in biological samples generally lead to inhomogeneous line broadening in 1D PRESS spectra, even severe overlapping of spectral peaks. By separating chemical shifts and J couplings into two different frequency dimensions, the localized two-dimensional (2D) J-resolved spectroscopy (JPRESS) is designed by adding an indirect spectral dimension in the original 1D PRESS to alleviate the spectral congestion [9, 10]. However, the JPRESS method remains sensitive to field inhomogeneity caused by macroscopic magnetic susceptibility variations in biological samples, especially in the investigations of large voxels that include different components. Although inhomogeneous line broadening in the J coupling dimension (F1) can be refocused by the spin-echo scheme [11], the overlapping of neighboring resonances in the chemical shift dimension (F2) makes it difficult to obtain exact J coupling information.

Many field shimming methods have been proposed to alleviate the influence of field inhomogeneity [12, 13]. However, the field inhomogeneity in biological tissues is generally hard to eliminate by conventional shimming methods. The voxel shimming approach has been used on MRI scanners [14]. This approach is time-consuming and is not suitable for biological samples when large detection volume is concerned. The magic angle spinning (MAS) technique [15–17] provides a feasible way to remove the influence of macroscopic magnetic susceptibility variations in biological samples by fast spinning [18, 19]. In general, the MAS technique requires a specialized probe suitable for typical NMR spectrometers and is not available for MRI scanners with broad inner bores. Furthermore, fragile organic textures, such as fish eggs and viscera, cannot endure fast spinning [20]. Thus a great demand for high-resolution 2D MRS methods which can be easily adopted to standard MRI scanners for practical applications has arisen.

It has been proved that intermolecular multiple-quantum coherences (iMQCs), originating from the distant dipolar interaction among spins in different molecules, can be used to recover high-resolution NMR spectra from inhomogeneous fields [21–23]. Recently, a method (dubbed as iDQCJRES) based on intermolecular double-quantum coherences (iDQCs) was proposed to obtain high-resolution 2D J-resolved NMR spectra in inhomogeneous fields [24]. The capability of iDQCJRES has been tested on a common 500 MHz NMR spectrometer with samples packed in 5 mm NMR tubes. However, the feasibility of the iDQCJRES method for practical MRS applications on MRI scanners with broad inner bores and low magnetic field strength remains uncertain. In this study, a localized iDQCJRES method was investigated on a 7 T MRI scanner with different samples and different voxel sizes. Experimental results were compared with those acquired by the conventional JPRESS method.

Theory

A PRESS like module [25], consisting of three slice-selective refocusing π RF pulses and corresponding slice-selective gradients along orthogonal directions, is integrated into the iDQCJRES sequence for spatial localization (Fig 1). In this localized iDQCJRES sequence, the PRESS like module can not only select the region of interest in the detected sample, but also refocus the resulting iDQC signals. The last slice-selective refocusing π RF pulse in the PRESS like module is inserted into the middle of the delay interval (2Δ) to preserve the desired signals before the distant dipolar interaction takes effect. Therefore, the non-selective π RF pulse used in the non-localized iDQCJRES can be omitted in the localized iDQCJRES sequence. Water suppression is a prerequisite for measurements of biological samples. Different from water suppression modules used in the original PRESS and JPRESS sequences, the double gradient echo W5 module implemented right before acquisition period is used in the localized iDQCJRES sequence to suppress the water signal [26, 27]. In this water suppression module, the crusher gradients are applied along the x, y and z directions. The first π/2 RF pulse is non-selective, and the second RF pulse (π/2)^I is selective for the water proton. A pair of linear coherence selection gradients (CSGs) with an area ratio of 1:-2 are employed along the z direction to select the desired coherence transfer pathway 0 → +2 → +1 → −1. Two indirect evolution periods, t₁ and t₂, are used for the desired signal evolution. Consider a homogeneous solution consisting of I (corresponding to solvent) and S (corresponding to solute) components, where I is an isolated single spin-1/2 system and S is an AX spin-1/2 system that includes S_k and S_l spins coupled by a J_kl scalar interaction. The evolution of two-spin order term for the desired signal from the localized iDQCJRES sequence can be understood intuitively by the product operator analysis as following, (1) where D_ISI_zS_z represents the distant dipolar interaction for iDQC between solvent and solute spins. According to the iMQC treatment [28], high temperature approximation is abandoned and the two spin term I_zS_z is the start point for the signal evolution. The localized iDQCJRES sequence starts with a non-selective π/2 RF pulse, and the iDQC term I⁺S⁺ is selected by the CSGs and evolved during the first evolution periods t₁/2. After that, the second (π/2)^I RF pulse selective for I spin transforms I⁺ into (0.5I⁺ − 0.5I⁻ − I_z) and only the term I_zS⁺ is persevered by the CSGs. Then the PRESS like module localizes the region of interest in the sample and overturns the coherence order from I_zS⁺ to I_zS⁻. Finally, the spin term I_zS⁻ evolves into observable signal by the distant dipolar interaction during the evolution period t₁/2+t₂/2+t₃. In this sequence, two indirect evolution periods, t₁ and t₂, are used and each is divided into two equal parts for the desired signal evolution. For the indirect evolution period t₁, I⁺S⁺ (iDQC term) evolution is involved in the first t₁/2 and S¯ evolution is involved in the second t₁/2, thus only the field inhomogeneity and J coupling are preserved in the F1 dimension. For the indirect evolution period t₂, I_zS⁺ evolution is involved in the first t₂/2 and S¯ evolution is involved in the second t₂/2, thus only the J coupling is observed in the F2 dimension. Before the acquisition period t₃, the distant dipolar interaction takes effects and transfers I_zS¯ into observable S¯ for signal acquisition. The field inhomogeneity effect remains in the F3 dimension. Since the double gradient echo W5 module right before acquisition only acts to suppress the water signal and does not influence the desired coherence transfer pathway and the desired solute signals, we ignore it in the theoretical analysis.

Download:

Fig 1. Pulse sequence diagram of the localized iDQCJRES.

Full vertical bar is the non-selective RF pulse, Gauss-shaped pulse is solvent-selective RF pulse, Sinc-shaped pulses stand for three slice-selective refocusing π RF pulses, trapezoids along three orthogonal directions are slice-selective gradients, vertical-line represent “W5” binomial π pulses. G and -2G are coherence selection gradients, G₁ and G₂ are crasher gradients for the water suppression. The coherence transfer pathway is presented and the product operators are applied to show the coherence states of solvent I and solute S spins.

https://doi.org/10.1371/journal.pone.0134109.g001

According to the previous report [24], the observable transverse magnetization of the S_k spin from the localized iDQCJRES sequence in an inhomogeneous field is given as (2) where ω_m is the frequency offset of spin m (m = I, S_k, S_l) in the rotating frame free of field inhomogeneity, and ΔB_m(r) is the inhomogeneous deviation of the magnetic field at the location of a particular m spin; and are equilibrium magnetizations per unit volume of S and I spins, respectively; μ₀ is the vacuum magnetic permeability, and γ is the gyromagnetic ratio. Eq (2) provides a quantitative expression of the 3D iDQCJRES signal between the solute spin S_k and the solvent spin I. In this equation, the terms represent the J coupling information along the F2 dimension, and the terms represent the evolution information of the solute along the F3 dimension, including chemical shift, field inhomogeneity, and J coupling. The terms are the iDQC terms along the F1 dimension, including chemical shift and field inhomogeneity of the solvent, and J coupling of the solute. If the spectrometer reference frequency coincides with the resonance frequency of I spin in B₀, i.e. ω_I = 0, the 3D iDQCJRES signal will split into two peaks and locate at and . In the localized iDQCJRES sequence, the dipolar correlation distance between S and I spins is inversely proportional to the area of CSGs, that is d_c = π/(γGδ) [29]. Generally, this distance is much smaller than the sample size, thus the magnetic field within the distance between S and I spins only varies slightly, and ΔB_I(r) is considered to be equal to . A shearing process on the F1-F3 plane is carried out to remove inhomogeneous line broadening along the F3 dimension. The frequency location for the signal in the sheared 3D spectrum becomes and . A projection of the sheared 3D spectrum onto the F2-F3 plane produces a desired high-resolution 2D J-resolved spectrum, and the signal locates at and . A clockwise rotation of this 2D iDQCJRES projection spectrum along F2 = 0 can separate chemical shifts and J couplings, resulting in peak positions at and , the same as the signal observed in a conventional 2D JPRESS spectrum.

Methods and Materials

All experiments were executed at 293 K using a Varian (Palo Alto, CA, USA) 7 T small animal magnetic resonance scanner with a 160 mm inner bore diameter and a 63/95 mm quad birdcage coil. The scanner was equipped with a gradient coil system producing a maximum gradient strength of 40 G/cm. The quad birdcage coil was well tuned to preserve high signal sensitivity. For comparison, the JPRESS sequence [30] for localized 2D J-resolved spectra was utilized as a reference scheme in our experiments on aqueous solution, pig brain tissue, and fish. For the localized iDQCJRES experiments, a 4-step phase cycling was applied: the phases for the first π/2 pulse, the second (π/2)^I RF pulse, and the receiver were (x, y,-x,-y), (x, x,-x,-x), and (x,-x,-x, x), respectively. The methods and experiments on biological samples were carried out in accordance with the approved guidelines. All biological samples (a whole fish and an intact pig brain tissue) used in our experiments were approved by the Institutional Review Board at Xiamen University, Xiamen, China.

Aqueous Solution

A γ-aminobutyric acid (GABA) aqueous solution (250 mM) filled in a plastic bottle with a volume of 68 cm³ was used to demonstrate the feasibility of the localized iDQCJRES sequence on the MRI scanner. Prior to spectral experiments, fast spin-echo images on coronal and axial orientations of the plastic bottle were acquired to show the localized regions. The magnetic field was deliberately degraded by altering the Z1 shimming coil current to produce broad peaks. The full width at half maximum (FWHM) of the water peak at 4.80 ppm was 180 Hz, and the full width at 10% maximum was 535 Hz. In this inhomogeneous field, the JPRESS and localized iDQCJRES sequences were applied to the same voxel with a size of 18 × 18 × 18 mm³. In addition, the magnetic field was shimmed using the standard shimming procedure provided in the MRI scanner, and then a JPRESS experiment and a localized iDQCJRES experiment on a voxel size of 6 × 6 × 6 mm³ under this shimmed field were performed for comparison. For the localized iDQCJRES experiments, the width of the π/2 hard RF pulse was 90 μs, the solvent-selective (π/2)^I RF pulse had a Gaussian shape with a width of 6.0 ms. The power levels of the π/2 hard RF pulse and solvent-selective π/2 pulse were 200 W and 250 mW, respectively. The parameters for the crusher gradients in the water suppression module were G₁ = 9.6 G/cm, G₂ = 26.9 G/cm, and δ′ = 3.0 ms. The width of sinc-shaped π pulses was 2.0 ms, and the parameters for the three slice-selective gradients were set to G_x = G_y = G_z = 0.29 G/cm with a duration of 2.0 ms for the 18 × 18 × 18 mm³ voxel, and G_x = G_y = G_z = 0.90 G/cm with a duration of 2.0 ms for the 6 × 6 × 6 mm³ voxel. Other parameters for the localized iDQCJRES experiments were as follows: the pulse repetition time TR = 2.0 s, the echo time (TE) 2Δ = 54 ms, the acquisition time = 100 ms, the average number = 4, and 10 × 30 × 600 points were acquired with spectral widths of 100 Hz × 50 Hz × 3000 Hz (F1 × F2 × F3) in 40 min. The localized iDQCJRES 3D data were processed using our custom-written program on MATLAB 7.11. For JPRESS experiments, the parameters for the spatial localization were set to G_x = 1.27 G/cm with a duration of 1.0 ms, G_y = G_z = 0.29 G/cm with a duration of 2.0 ms for the 18 × 18 × 18 mm³ voxel, and G_x = 3.82 G/cm with a duration of 1.0 ms, G_y = G_z = 0.90 G/cm with a duration of 2.0 ms for the 6 × 6 × 6 mm³ voxel. The variable power and optimized relaxation delays (VAPOR) module was used for water suppression. The TR/TE was 2000/15 ms, the acquisition time was 100 ms, the average number was 4, and 30 × 600 points were acquired with spectral widths of 50 Hz × 3000 Hz (F1 × F2) in 4 min.

Pig Brain Tissue

A sample of intact pig brain tissue was applied to show the capability of the localized iDQCJRES sequence on biological tissues. The pig brain tissue was purchased from a local retailer named Fujian New Hua Du Supermarket Co., LTD. (24°44'N, 118°09'E), and the item number for this sample was FJNQ896563328. The pig brain tissue was carefully packed in a fresh bag and kept in a fresh layer of fridge at 5°C for about one hour before the experiments. Prior to spectral measurements, fast spin-echo images on axial and coronal orientations were acquired with TR/TE = 2500/40 ms and imaging matrix = 256 × 256 in circa 5 min. The iDQCJRES experiment was performed without any field shimming. In this experiment, the widths of π/2 hard pulse and solvent-selective (π/2)^I RF pulse were 98 μs and 5.9 ms, respectively. The parameters of the CSGs and the WS module were the same as those used for the GABA aqueous solution experiment. The parameters of spatial localization were G_x = G_y = G_z = 0.29 G/cm with a duration of 2.0 ms for the 18 × 18 × 18 mm³ voxel. The TR/TE was 2000/36 ms, the acquisition time was 105 ms, the average number was 16, and 9 × 30 × 650 points were acquired with spectral widths of 100 Hz × 50 Hz × 3000 Hz (F1 × F2 × F3) in 144 min. A JPRESS experiment on the same voxel under the same field condition was performed for comparison. As a reference, a JPRESS experiment with a small voxel of 6 × 6 × 6 mm³ under a relatively homogeneous field was performed after the standard shimming procedure. The VAPOR module was used for water suppression. The JPRESS experiments were acquired with TR/TE = 2000/15 ms, 16 average number, 30 × 650 points for spectral widths of 50 Hz × 3000 Hz (F1 × F2) in 16 min.

A Whole Fish

To show the applicability of the localized iDQCJRES sequence on real biological samples with integrated organism, we performed a postmortem study on a whole fish (Harpadon nehereus). The fish sample was purchased from the same local retailer where we bought the pig brain tissue, with an item number of FJDS042108187. This fish sample was originally supplied by Dongshan fishery (23°33'N, 117°17'E). The purchased fish was carefully packed in a fresh bag and preserved in a fresh layer of fridge at 5°C for about one hour before experiments. The standard shimming procedure was executed to optimize the field homogeneity. A fast spin-echo MRI experiment was carried out to display inner structures of the fish in axial and coronal planes. For the iDQCJRES experiment, the width of the π/2 hard pulse was 111 μs and the width of solvent-selective (π/2)^I RF pulse was 6.2 ms. The parameters of the CSGs and the WS module were the same as those used in the aqueous solution experiment. The parameters of spatial localization were G_x = G_y = G_z = 0.33 G/cm with a duration of 2.0 ms for the 16 × 16 × 16 mm³ voxel. The TR/TE was 2000/36 ms, the acquisition time was 120 ms, the average number was 32, and 9 × 25 × 720 points were acquired with spectral widths of 100 Hz × 50 Hz × 3000 Hz (F1 × F2 × F3) in 240 min. Two JPRESS experiments on the voxels of 16 × 16 × 16 mm³ and 6 × 6 × 6 mm³ were performed, respectively, with TR/TE = 2000/15 ms, 32 average number, and 25 × 720 points for spectral widths of 50 Hz × 3000 Hz (F1 × F2) in 26.7 min.

Results and Discussion

The feasibility of the localized iDQCJRES on recovering high-resolution 2D J-resolved spectra from inhomogeneous fields at the 7 T MRI scanner is verified by the GABA aqueous solution experiment (Fig 2). The fast spin-echo images of the GABA aqueous solution filled in a plastic bottle at coronal and axial sections are given to show the localized regions (Fig 2A). After a clockwise rotation of 45°, the 2D JPRESS spectrum acquired from the voxel of 6 × 6 × 6 mm³ after the standard shimming procedure is presented as a reference (Fig 2B). Owing to the broad inner bore of the 7 T MRI scanner, it is hard to keep the magnetic field absolutely homogeneous. Thus the FWHM of the peak at 2.3 ppm along the F2 dimension of the 2D JPRESS spectrum remains 32 Hz (Fig 2B). The 2D J-resolved information can be obtained, and three coupled peaks of GABA are observed along the F2 dimension, and the related J coupling constants and multiplet patterns are presented in the F1 dimension. A 2D J-resolved spectrum obtained from the localized iDQCJRES method with the same voxel size and field condition is also presented (Fig 2C). Benefiting from the immunity to field inhomogeneity of the iDQCJRES method, the FWHM of the peak at 2.3 ppm along the F2 dimension of the 2D localized iDQCJRES spectrum reaches to 18 Hz. It is obvious that the signal to noise ratio (SNR) of this spectrum is lower than that of the conventional JPRESS spectrum (Fig 2B). To make a clear comparison, we performed SNR calculations on these two experiments. The SNR is calculated by dividing the intensity of the peak at 3.01 ppm by the standard deviation of noise signals in the region between 5.0 and 5.5 ppm along the 1D projection [31]. The SNR is 15.3 for Fig R2C, while it is 114.1 for Fig R2B. So the SNR of the 2D localized iDQCJRES spectrum only presents 13.4% of that of the conventional 2D JPRESS spectrum.

Download:

Fig 2. Results of the GABA aqueous solution.

(A) Spin echo images of the GABA solution filled into a plastic bottle. A large voxel of 18 × 18 × 18 mm³ is marked by a green rectangle, and a small voxel of 6 × 6 × 6 mm³ is marked by a red rectangle. (B) Conventional 2D JPRESS spectrum after a clockwise rotation of 45° and its projection along the F2 axis acquired from the small voxel in a relatively homogeneous field after the field shimming procedure. (C) Localized iDQCJRES spectrum and its 1D J-decoupled projection along the F3 axis acquired from the small voxel under the same field homogeneity as (B). (D) Conventional 1D non-localized spectrum acquired in an inhomogeneous field with a 180 Hz FWHM of the water peak at 4.8 ppm. (E) Conventional 2D JPRESS spectrum after a clockwise rotation of 45° and its projection along the F2 axis acquired from the large voxel in an inhomogeneous field. (F) Localized iDQCJRES spectrum and its 1D J-decoupled projection along the F3 axis acquired from the large voxel in the same inhomogeneous field as (E).

https://doi.org/10.1371/journal.pone.0134109.g002

However, when the magnetic field is deliberately deshimmed, the valuable spectral information will be lost in conventional spectra. A conventional 1D non-localized spectrum of the sample was acquired without water suppression to show the field condition (Fig 2D). The FWHM of the water peak at 4.8 ppm is 180 Hz, and the information of chemical shifts and J couplings is erased by inhomogeneous line broadening. Similarly, the conventional JPRESS spectrum also suffers from the influence of field inhomogeneity. It is difficult to extract satisfactory information from the 2D JPRESS spectrum acquired from the voxel of 18 × 18 × 18 mm³ under this inhomogeneous field (Fig 2E). All signal peaks stretch along the F2 axis due to inhomogeneous line broadening. Although the field inhomogeneity can be refocused by spin echo scheme along the F1 dimension, the overlapping among neighboring peaks obscures J coupling measurement. However, under the same inhomogeneous field and from the same voxel, a high-resolution 2D J-resolved spectrum can be obtained by using the localized iDQCJRES method (Fig 2F). Compared to the conventional 2D JPRESS spectrum (Fig 2E), the spectral resolution along the chemical shift dimension (F3) is significantly improved in the 2D localized iDQCJRES spectrum, and the FWHM for the peak at 2.3 ppm is reduced to 30 Hz. In addition, J coupling constants and multiplet patterns are explicitly shown in the F2 dimension. The spectral features provided in the 2D localized iDQCJRES spectrum in the inhomogeneous field are the same as those obtained from the conventional 2D JPRESS spectrum in the relatively homogeneous field. Furthermore, the SNR of the 2D iDQCJRES spectrum acquired from the relatively large voxel of 18 × 18 × 18 mm³ in the inhomogeneous field is 106.5 (Fig 2F), close to the SNR of the conventional 2D JPRESS spectrum acquired from the voxel of 6 × 6 × 6 mm³ under the relatively homogeneous field (Fig 2B). Thus, a relatively large voxel can partially compensate for the weakness of the localized iDQCJRES method on SNR. It is notable that the echo time used in the localized iDQCJRES experiments is longer than that used in the conventional JPRESS experiments (54 ms for iDQCJRES, 15 ms for JPRESS). Long echo time for the conventional JPRESS will lead to signal decay caused by the transverse relaxation (T₂ relaxation). Therefore, the default echo time of 15 ms on the MRI scanner was used in the JPRESS experiments to preserve maximal JPRESS signals. In the case of iMQC MRS experiments, since the distant dipolar interaction needs some time to take effect (the so-called “demagnetizing time”), the signal grows first and then decays following T₂ relaxation [29]. We performed an arrayed experiment to seek optimal echo time for maximal iDQCJRES signal. It turns out that the echo time of 54 ms is optimal for the iDQCJRES experiments.

The experimental results of the intact pig brain tissue are presented in Fig 3. The fast spin-echo images of the brain tissue at coronal and axial sections are given to show voxel positions (Fig 3A). Due to the field inhomogeneity caused by intrinsic macroscopic magnetic susceptibility variations, hardly any spectral information can be obtained from the conventional 1D non-localized spectrum (Fig 3B). The conventional 2D JPRESS spectrum and its 1D projection along the F2 dimension acquired from the voxel of 6 × 6 × 6 mm³ are shown in Fig 3C. In MRS studies of biological tissues, the field inhomogeneity is directly dependent on the selected voxel size [32]. Therefore the field homogeneity in the JPRESS experiment on a small voxel of 6 × 6 × 6 mm³ can be well after the field shimming procedure. The FWHM of choline (Cho) at 3.20 ppm is 39 Hz (Fig 3C) and the major metabolite peaks are observed and assigned. The J coupling information of some metabolites can be measured, such as D-lactic acid (Lac) at 1.34 ppm and alanine (Ala) at 1.53 ppm. However, when a relatively large voxel of 18 × 18 × 18 mm³ is selected, the field homogeneity decreases remarkably due to the variations of magnetic susceptibility among various structures of the brain tissue. The resulting 2D JPRESS spectrum is influenced by inhomogeneous line broadening and the desired spectral information for metabolite analyses is lost (Fig 3D). The peaks are overlapped along the F2 dimension and the J coupling information is hard to identify along the F1 dimension. Although a large voxel is beneficial to signal intensity, the aggravated field inhomogeneity makes the application of the conventional JPRESS method on large voxels challenging.

Download:

Fig 3. Results of an intact pig brain tissue.

(A) Spin echo images of the sample. A large voxel of 18 × 18 × 18 mm³ is marked by a green dashed rectangle, and a small voxel of 6 × 6 × 6 mm³ is marked by a red dashed rectangle. (B) Conventional 1D non-localized spectrum and the expanded region for metabolites. (C) Conventional 2D JPRESS spectrum after a clockwise rotation of 45° and its projection along the F2 axis acquired from the small voxel after the field shimming procedure. (D) Conventional 2D JPRESS spectrum after a clockwise rotation of 45° and its projection along the F2 axis acquired from the large voxel without the field shimming procedure. (E) 2D localized iDQCJRES spectrum and its 1D J-decoupled projection along the F3 axis acquired from the large voxel without the field shimming procedure.

https://doi.org/10.1371/journal.pone.0134109.g003

The localized 2D iDQCJRES spectrum acquired from the large voxel of 18 × 18 × 18 mm³ without any field shimming procedure is given in Fig 3E. Compared to the conventional 2D JPRESS spectrum, the spectral resolution is significantly improved. Taking the Cho peak at 3.18 ppm for an example, its FWHM is 30 Hz in this localized iDQCJRES spectrum (Fig 3E), 100 Hz in the conventional 2D JPRESS spectrum of the voxel of 18 × 18 × 18 mm³ without the field shimming (Fig 3C), and 39 Hz in the conventional 2D JPRESS spectrum of the voxel of 6 × 6 × 6 mm³ after the field shimming (Fig 3D). Clearly, the localized iDQCJRES spectrum can yield satisfactory spectral information, and this information is even better than that provided in the conventional JPRESS spectrum acquired from the small voxel in the shimmed field. A considerable number of metabolite signals are resolved and assigned [33] in the localized iDQCJRES spectrum. To make a comparison of the results obtained by the localized iDQCJRES method from the large voxel and by the conventional JPRESS method from the small voxel, we list the ¹H chemical shifts, multiplet patterns, and J coupling constants extracted from the two spectra (Fig 3C and 3E) in Table 1. Twelve peaks could be assigned to 11 metabolites in the localized iDQCJRES spectrum from the large voxel, while 11 peaks could be assigned to 10 metabolites in the conventional JPRESS spectrum from the small voxel.

Download:

Table 1. Comparison of spectral results of an intact pig brain tissue obtained by localized iDQCJRES and conventional JPRESS methods.

https://doi.org/10.1371/journal.pone.0134109.t001

In a previous study, a localized MRS method based on intermolecular single-quantum coherences has been used to obtain 1D spectrum with enhanced resolution on pig brain tissues [34]. The spectral resolution therein was not enough for observing J coupling splitting, and only chemical shifts could be observed. In the localized iDQCJRES spectrum, chemical shifts and J couplings are provided along two separate dimensions. Chemical shifts directly point to metabolite assignments and J couplings aid metabolite identification. It can be noticed that the apparent signal intensity of Lac is decreased in the localized iDQCJRES spectrum (Fig 3E) compared to that in the conventional JPRESS spectrum (Fig 3D). Similar result was observed in the previous MRS study on the brain tissue [32, 34, 35]. The main reason is that the lipid signal at 1.25 ppm generally overlaps with the Lac signal at 1.30 ppm due to the insufficient spectral resolution. The contribution of lipid signal to the signal intensity can be observed in the JPRESS spectrum with a short echo time. Because a long echo time was adopted in the iDQCJRES experiment, the decay of lipid signal became severe due to its short transverse relaxation time, hence a relatively weak Lac signal free of the interference of lipid signal was obtained in the iDQCJRES spectrum.

The ability of the localized iDQCJRES method on enhancing spectral resolution from strongly structured biological tissues is exhibited by the experiments of a whole fish (Harpadon nehereus) (Fig 4). The coronal and axial spin-echo images of the fish are displayed in Fig 4A. The length of the fish is about 200 mm and only part of the axial image is given. A large voxel of 16 × 16 × 16 mm³ containing the fish spinal cord is marked by a green dashed box in the two images, while a small voxel of 6 × 6 × 6 mm³ containing only the fish flesh is marked by a red dashed box in these images. Hardly any spectral information can be obtained from the conventional 1D non-localized spectrum due to the field inhomogeneity (Fig 4B). When the conventional JPRESS method is applied to the small voxel, the field inhomogeneity can be partially removed by field shimming, and a 2D JPRESS spectrum with acceptable resolution can be obtained (Fig 4C). The FWHM of Cho at 3.22 ppm is 38 Hz and some metabolites can be observed. The J coupling information of the metabolites, such as the methyl group of low-density lipoprotein (LDL) at 0.93 ppm and lactate (Lac) at 1.34 ppm, can be extracted along the F1 dimension. Because of the prolate shapes and surrounding bone structures, the field shimming for the fish spinal cord region is generally challenging [36]. Thus, when the large voxel containing the fish spinal cord is selected, the quality of the resulting 2D JPRESS spectrum remarkably decreases even after field shimming (Fig 4D). The FWHM of the water peak at 4.80 pm is 120 Hz. Most metabolite peaks are overlapped and lost along the F2 dimension, and only the J coupling splitting of the methyl group of low-density lipoprotein (LDL) at 0.75 ppm can be observed along the F1 dimension.

Download:

Fig 4. Results of a whole fish (Harpadon nehereus).

(A) Spin echo images of the fish. A large voxel of 16 × 16 × 16 mm³ containing the fish spinal cord is marked by a green dashed box, and a small voxel of 6 × 6 × 6 mm³ containing the fish tissue is marked by a red dashed box. (B) Conventional 1D non-localized spectrum and its expanded region for metabolites. (C) Conventional 2D JPRESS spectrum after a clockwise rotation of 45° and its projection along the F2 axis acquired from the small voxel after the field shimming procedure. (D) Conventional 2D JPRESS spectrum after a clockwise rotation of 45° and its projection along the F2 axis acquired from the large voxel after the field shimming procedure. (E) 2D localized iDQCJRES spectrum and its 1D J-decoupled projection along the F3 axis acquired from the large voxel without the field shimming procedure.

https://doi.org/10.1371/journal.pone.0134109.g004

A 2D localized iDQCJRES spectrum, acquired from the voxel of 16 × 16 × 16 mm³ containing the fish spinal cord and without field shimming, provides 2D J-resolved information with enhanced spectral resolution (Fig 4E). The FWHM of Cho at 3.25 ppm is 28 Hz along the F3 dimension in this 2D spectrum, better than 38 Hz along the F2 dimension of the 2D JPRESS spectrum acquired from the small voxel after field shimming. A considerable number of metabolite signals are identified and assigned. Benefiting from the spectral resolution enhancement, some weak metabolite signals invisible in the JPRESS spectrum (Fig 4D) can be observed in the iDQCJRES spectrum (Fig 4E), such as alanine (Ala) at 1.48 ppm. The assignment of the observed peaks according to literature [20, 37] is given in Fig 4. To make a clear comparison between the results obtained by the iDQCJRES method from the large voxel without field shimming and by the conventional JPRESS method from the small voxel after field shimming, we list the ¹H chemical shifts, multiplet patterns, and J coupling constants in Table 2. Fourteen peaks are assigned to 11 metabolites in the localized iDQCJRES spectrum, while 7 peaks are assigned to 6 metabolites in the conventional JPRESS spectrum. Note that the observed peaks in the spectra acquired from the same fish with different methods have some differences. For example, glutamate/glutamine (Glu/Gln) at 3.75 ppm and glycine at 3.55 ppm are present in the iDQCJRES spectrum while absent in the conventional JPRESS spectrum. This may be attributed to the complex circumstance, magnetic susceptibility gradient between the fish tissue and the air in abdominal cavity, and intrinsic magnetic susceptibility variations in fish itself among muscle tissues, viscera and bones. The difference in the voxel selection may pose another possible reason.

Download:

Table 2. Comparison of 1H NMR spectral results of a whole fish (Harpadon nehereus) obtained using iDQCJRES and conventional JPRESS methods.

https://doi.org/10.1371/journal.pone.0134109.t002

All above results show that the localized iDQCJRES method can be applied for direct measurement of biological samples to obtain high-resolution J-resolved information, without the limitation of voxel size and field shimming requirement. However, the localized iDQCJRES method also has disadvantages in signal sensitivity and experimental time in contrast to the conventional JPRESS method. On the aspect of signal sensitivity, the use of large voxel can partially compensate for the low signal intensity. Besides, high sensitivity probes [38] and parallel coils [39] on MRI scanners may be useful for the improvement of signal intensity for the localized iDQCJRES method. On the aspect of acquisition time, the localized iDQCJRES experiment takes longer acquisition time than the conventional JPRESS since 3D acquisition is required. The spatial encoding scheme [40] may be applied to the t₁ period of the localized iDQCJRES sequence, so t₁ increments can be sampled in a single scan, and the acquisition time can be shortened to the level of the conventional 2D JPRESS sequence. It is obvious that both of the conventional JPRESS and the localized iDQCJRES method have their own advantages and drawbacks in practical applications. The conventional JPRESS method is useful for the measurements of metabolites in a specific small area of interest under a relatively homogeneous magnetic field. The localized iDQCJRES method is more useful for the measurements of metabolites in a relatively large area with field inhomogeneity, such as the lesion area in a large animal or human body [41]. In practical applications, there is no technique applicable to all circumstances, and the localized iDQCJRES method may provide a complementary way to the conventional JPRESS method for MRS measurements of biological samples on MRI systems.

Conclusion

In this work, the implementation of the localized iDQCJRES method on a 7 T MRI scanner is studied. The experiment on a GABA aqueous solution reveals the feasibility of the localized iDQCJRES method on refocusing inhomogeneous line broadening on the 7 T MRI scanner. Spatially localized applications on biological samples are demonstrated on an intact pig brain tissue and a whole fish. The spectral information observed in the localized iDQCJRES spectra acquired under inhomogeneous fields (due to large voxel and no field shimming) is similar to that provided by the JPRESS spectra acquired in relatively homogeneous fields (due to small voxel and field shimming). Our experimental observations clearly illuminate the advantages of the localized iDQCJRES method for 2D MRS applications on MRI systems with broad inner bores. This method presents an alternative to the conventional JPRESS method for MRS measurements on biological samples, and may be a promising tool for in vivo MRS applications.

Acknowledgments

The authors are grateful for the technical assistance from Hao Chen, Kaiyu Wang, and Shuai Chen.

Author Contributions

Conceived and designed the experiments: CT YH. Performed the experiments: CT YH. Analyzed the data: CT YH. Contributed reagents/materials/analysis tools: SC YH CT. Wrote the paper: CT YH SC. No.

References

1. Kurhanewicz J, Swanson MG, Nelson SJ, Vigneron DB. Combined magnetic resonance imaging and spectroscopic imaging approach to molecular imaging of prostate cancer. J Magn Reson Imaging. 2002; 16(4): 451–463. pmid:12353259
- View Article
- PubMed/NCBI
- Google Scholar
2. Shah T, Jayasundar R, Singh VP, Sarkar C. In vivo MRS study of intraventricular tumors. J Magn Reson Imaging. 2011; 34(5): 1053–1059. pmid:22002756
- View Article
- PubMed/NCBI
- Google Scholar
3. Tzeng SR, Kalodimos CG. Protein activity regulation by conformational entropy. Nature. 2012; 488(7410): 236–240. pmid:22801505
- View Article
- PubMed/NCBI
- Google Scholar
4. Nath N, Lokesh , Suryaprakash N. Measurement and applications of long-range heteronuclear scalar couplings: Recent experimental and theoretical developments. ChemPhysChem. 2012; 13(3): 645–660. pmid:22302693
- View Article
- PubMed/NCBI
- Google Scholar
5. Wu EL, Engstrom O, Jo S, Stuhlsatz D, Yeom MS, Klauda JB, et al. Molecular dynamics and NMR spectroscopy studies of E. coli lipopolysaccharide structure and dynamics. Biophys J. 2013; 105(6): 1444–1455. pmid:24047996
- View Article
- PubMed/NCBI
- Google Scholar
6. Huang RZ, Gao HC, Zhang L, Jia JM, Liu X, Zheng P, et al. Borna disease virus infection perturbs energy metabolites and amino acids in cultured human oligodendroglia cells. Plos One. 2012; 7(9): e44665. pmid:22970281
- View Article
- PubMed/NCBI
- Google Scholar
7. Solanky BS, Abdel-Aziz K, Yiannakas MC, Berry AM, Ciccarelli O, Wheeler-Kingshott CAM. In vivo magnetic resonance spectroscopy detection of combined glutamate-glutamine in healthy upper cervical cord at 3T. Nmr Biomed. 2013; 26(3): 357–366. pmid:23281170
- View Article
- PubMed/NCBI
- Google Scholar
8. Herminghaus S, Pilatus U, Moller-Hartmann W, Raab P, Lanfermann H, Schlote W, et al. Increased choline levels coincide with enhanced proliferative activity of human neuroepithelial brain tumors. Nmr Biomed. 2002; 15(6): 385–392. pmid:12357552
- View Article
- PubMed/NCBI
- Google Scholar
9. Bruch RC, Bruch MD. Two-dimensional J-resolved proton NMR spectroscopy of oligomannosidic glycopeptides. J Biol Chem. 1982; 257(7): 3409–3413. pmid:7061488
- View Article
- PubMed/NCBI
- Google Scholar
10. Pola A, Sadananthan SA, Yaligar J, Nagarajan V, Han WP, Kuchel PW, et al. Skeletal muscle lipid metabolism studied by advanced magnetic resonance spectroscopy. Prog Nucl Magn Reson Spectrosc. 2012; 65: 66–76. pmid:22781315
- View Article
- PubMed/NCBI
- Google Scholar
11. Ludwig C, Viant MR. Two-dimensional J-resolved NMR spectroscopy: Review of a key methodology in the metabolomics toolbox. Phytochem Anal. 2010; 21(1): 22–32. pmid:19904730
- View Article
- PubMed/NCBI
- Google Scholar
12. Terpstra M, Andersen PM, Gruetter R. Localized eddy current compensation using quantitative field mapping. J Magn Reson. 1998; 131(1): 139–143. pmid:9533916
- View Article
- PubMed/NCBI
- Google Scholar
13. Kim DH, Adalsteinsson E, Glover GH, Spielman DM. Regularized higher-order in vivo shimming. Magn Reson Med. 2002; 48(4): 715–722. pmid:12353290
- View Article
- PubMed/NCBI
- Google Scholar
14. Springer F, Machann J, Schwenzer NF, Ballweg V, Wurslin C, Schneider JH, et al. Quantitative assessment of intrahepatic lipids using fat-selective imaging with spectral-spatial excitation and in-/opposed-phase gradient echo imaging techniques within a study population of extremely obese patients feasibility on a short, wide-bore MR scanner. Invest Radiol. 2010; 45(8): 484–490. pmid:20479651
- View Article
- PubMed/NCBI
- Google Scholar
15. Tugnoli V, Schenetti L, Mucci A, Nocetti L, Toraci C, Mavilla L, et al. A comparison between in vivo and ex vivo HR-MAS ¹H MR spectra of a pediatric posterior fossa lesion. Int J Mol Med. 2005; 16(2): 301–307. pmid:16012766
- View Article
- PubMed/NCBI
- Google Scholar
16. Righi V, Durante C, Cocchi M, Calabrese C, Di Febo G, Lecce F, et al. Discrimination of healthy and neoplastic human colon tissues by ex vivo HR-MAS NMR spectroscopy and chemometric analyses. J Proteome Res. 2009; 8(4): 1859–1869. pmid:19714812
- View Article
- PubMed/NCBI
- Google Scholar
17. Hamaed H, Pawlowski JM, Cooper BFT, Fu RQ, Eichhorn SH, Schurko RW. Application of solid-state ³⁵Cl NMR to the structural characterization of hydrochloride pharmaceuticals and their polymorphs. J Am Chem Soc. 2008; 130(33): 11056–11065. pmid:18656917
- View Article
- PubMed/NCBI
- Google Scholar
18. Sitter B, Sonnewald U, Spraul M, Fjosne HE, Gribbestad IS. High-resolution magic angle spinning MRS of breast cancer tissue. Nmr Biomed. 2002; 15(5): 327–337. pmid:12203224
- View Article
- PubMed/NCBI
- Google Scholar
19. Nestor G, Bankefors J, Schlechtriem C, Brannas E, Pickova J, Sandstrom C. High-resolution H-1 magic angle spinning NMR spectroscopy of intact arctic char (salvelinus alpinus) muscle. quantitative analysis of n-3 fatty acids, EPA and DHA. J Agr Food Chem. 2010; 58(20): 10799–10803.
- View Article
- Google Scholar
20. Cai HH, Chen YS, Cui XH, Cai SH, Chen Z. High-Resolution H-1 NMR spectroscopy of fish muscle, eggs and small whole fish via hadamard-encoded intermolecular multiple-quantum coherence. Plos One. 2014; 9(1): e86422. pmid:24466083
- View Article
- PubMed/NCBI
- Google Scholar
21. Vathyam S, Lee S, Warren WS. Homogeneous NMR spectra in inhomogeneous fields. Science. 1996; 272(5258): 92–96. pmid:8600541
- View Article
- PubMed/NCBI
- Google Scholar
22. Lin YY, Ahn S, Murali N, Brey W, Bowers CR, Warren WS. High-resolution, > 1 GHz NMR in unstable magnetic fields. Phys Rev Lett. 2000; 85(17): 3732–3735. pmid:11030993
- View Article
- PubMed/NCBI
- Google Scholar
23. Chen Z, Chen ZW, Zhong JH. High-resolution NMR spectra in inhomogeneous fields via IDEAL (intermolecular dipolar-interaction enhanced all lines) method. J Am Chem Soc. 2004; 126(2): 446–447. pmid:14719924
- View Article
- PubMed/NCBI
- Google Scholar
24. Huang YQ, Cal SH, Zhang ZY, Chen Z. High-resolution two-dimensional J-resolved NMR spectroscopy for biological systems. Biophys J. 2014; 106(9): 2061–2070. pmid:24806938
- View Article
- PubMed/NCBI
- Google Scholar
25. Bottomley PA. Spatial localization in NMR-spectroscopy in vivo. Ann Ny Acad Sci. 1987; 508: 333–348. pmid:3326459
- View Article
- PubMed/NCBI
- Google Scholar
26. Liu ML, Mao XA, Ye CH, Huang H, Nicholson JK, Lindon JC. Improved WATERGATE pulse sequences for solvent suppression in NMR spectroscopy. J Magn Reson. 1998; 132(1): 125–129.
- View Article
- Google Scholar
27. Zheng G, Price WS. Solvent signal suppression in NMR. Prog Nucl Magn Reson Spectrosc. 2010; 56(3): 267–288. pmid:20633355
- View Article
- PubMed/NCBI
- Google Scholar
28. Ahn S, Warren WS, Lee S. Quantum treatment of intermolecular multiple-quantum coherences with intramolecular J coupling in solution NMR. J Magn Reson. 1997; 128(2): 114–129. pmid:9356265
- View Article
- PubMed/NCBI
- Google Scholar
29. Lee S, Richter W, Vathyam S, Warren W. Quantum treatment of the effects of dipole–dipole interactions in liquid nuclear magnetic resonance. J Chem Phys. 1996; 105(3): 874–900.
- View Article
- Google Scholar
30. Thomas MA, Ryner LN, Mehta MP, Turski PA, Sorenson JA. Localized 2D J-resolved ¹H MR spectroscopy of human brain tumors in vivo. J Magn Reson Imaging. 1996; 6(3): 453–459. pmid:8724410
- View Article
- PubMed/NCBI
- Google Scholar
31. Ernst RR, Bodenhausen G, Wokaun A. Principles of nuclear magnetic resonance in one and two dimensions. Oxford: Clarendon Press; 1987.
32. Balla DZ, Melkus G, Faber C. Spatially localized intermolecular zero-quantum coherence spectroscopy for in vivo applications. Magn Reson Med. 2006; 56(4): 745–753. pmid:16897767
- View Article
- PubMed/NCBI
- Google Scholar
33. Govindaraju V, Young K, Maudsley AA. Proton NMR chemical shifts and coupling constants for brain metabolites. Nmr Biomed. 2000; 13(3): 129–153. pmid:10861994
- View Article
- PubMed/NCBI
- Google Scholar
34. Cui XH, Bao JF, Huang YQ, Cai SH, Chen Z. In vivo spatially localized high resolution ¹H MRS via intermolecular single-quantum coherence of rat brain at 7 T. J Magn Reson Imaging. 2013; 37(2): 359–364. pmid:23034817
- View Article
- PubMed/NCBI
- Google Scholar
35. Faber C. Solvent-localized NMR spectroscopy using the distant dipolar field: A method for NMR separations with a single gradient. J Magn Reson. 2005; 176(1): 120–124. pmid:15990343
- View Article
- PubMed/NCBI
- Google Scholar
36. Balla DZ, Faber C. Localized intermolecular zero-quantum coherence spectroscopy in vivo. Concept Magn Reson A. 2008; 32A(2): 117–133.
- View Article
- Google Scholar
37. Gribbestad IS, Aursand M, Martinez I. High-resolution ¹H magnetic resonance spectroscopy of whole fish, fillets and extracts of farmed Atlantic salmon (Salmo salar) for quality assessment and compositional analyses. Aquaculture. 2005; 250(1–2): 445–457.
- View Article
- Google Scholar
38. Sack M, Wetterling F, Sartorius A, Ende G, Weber-Fahr W. Signal-to-noise ratio of a mouse brain ¹³C CryoProbe^TM system in comparison with room temperature coils: Spectroscopic phantom and in vivo results. Nmr Biomed. 2014; 27(6): 709–715. pmid:24692120
- View Article
- PubMed/NCBI
- Google Scholar
39. Hall EL, Stephenson MC, Price D, Morris PG. Methodology for improved detection of low concentration metabolites in MRS: Optimised combination of signals from multi-element coil arrays. Neuroimage. 2014; 86: 35–42. pmid:23639258
- View Article
- PubMed/NCBI
- Google Scholar
40. Tal A, Frydman L. Single-scan multidimensional magnetic resonance. Prog Nucl Magn Reson Spectrosc. 2010; 57(3): 241–292. pmid:20667401
- View Article
- PubMed/NCBI
- Google Scholar
41. Träber F, Block W, Freymann N, Gür O, Kucinski T, Hammen T, et al. A multicenter reproducibility study of single-voxel ¹H-MRS of the medial temporal lobe. Eur Radiol. 2006; 16(5): 1096–1103. pmid:16416279
- View Article
- PubMed/NCBI
- Google Scholar

[ref1] 1. Kurhanewicz J, Swanson MG, Nelson SJ, Vigneron DB. Combined magnetic resonance imaging and spectroscopic imaging approach to molecular imaging of prostate cancer. J Magn Reson Imaging. 2002; 16(4): 451–463. pmid:12353259
View Article
PubMed/NCBI
Google Scholar

[2] View Article

[3] PubMed/NCBI

[4] Google Scholar

[ref2] 2. Shah T, Jayasundar R, Singh VP, Sarkar C. In vivo MRS study of intraventricular tumors. J Magn Reson Imaging. 2011; 34(5): 1053–1059. pmid:22002756
View Article
PubMed/NCBI
Google Scholar

[6] View Article

[7] PubMed/NCBI

[8] Google Scholar

[ref3] 3. Tzeng SR, Kalodimos CG. Protein activity regulation by conformational entropy. Nature. 2012; 488(7410): 236–240. pmid:22801505
View Article
PubMed/NCBI
Google Scholar

[10] View Article

[11] PubMed/NCBI

[12] Google Scholar

[ref4] 4. Nath N, Lokesh , Suryaprakash N. Measurement and applications of long-range heteronuclear scalar couplings: Recent experimental and theoretical developments. ChemPhysChem. 2012; 13(3): 645–660. pmid:22302693
View Article
PubMed/NCBI
Google Scholar

[14] View Article

[15] PubMed/NCBI

[16] Google Scholar

[ref5] 5. Wu EL, Engstrom O, Jo S, Stuhlsatz D, Yeom MS, Klauda JB, et al. Molecular dynamics and NMR spectroscopy studies of E. coli lipopolysaccharide structure and dynamics. Biophys J. 2013; 105(6): 1444–1455. pmid:24047996
View Article
PubMed/NCBI
Google Scholar

[18] View Article

[19] PubMed/NCBI

[20] Google Scholar

[ref6] 6. Huang RZ, Gao HC, Zhang L, Jia JM, Liu X, Zheng P, et al. Borna disease virus infection perturbs energy metabolites and amino acids in cultured human oligodendroglia cells. Plos One. 2012; 7(9): e44665. pmid:22970281
View Article
PubMed/NCBI
Google Scholar

[22] View Article

[23] PubMed/NCBI

[24] Google Scholar

[ref7] 7. Solanky BS, Abdel-Aziz K, Yiannakas MC, Berry AM, Ciccarelli O, Wheeler-Kingshott CAM. In vivo magnetic resonance spectroscopy detection of combined glutamate-glutamine in healthy upper cervical cord at 3T. Nmr Biomed. 2013; 26(3): 357–366. pmid:23281170
View Article
PubMed/NCBI
Google Scholar

[26] View Article

[27] PubMed/NCBI

[28] Google Scholar

[ref8] 8. Herminghaus S, Pilatus U, Moller-Hartmann W, Raab P, Lanfermann H, Schlote W, et al. Increased choline levels coincide with enhanced proliferative activity of human neuroepithelial brain tumors. Nmr Biomed. 2002; 15(6): 385–392. pmid:12357552
View Article
PubMed/NCBI
Google Scholar

[30] View Article

[31] PubMed/NCBI

[32] Google Scholar

[ref9] 9. Bruch RC, Bruch MD. Two-dimensional J-resolved proton NMR spectroscopy of oligomannosidic glycopeptides. J Biol Chem. 1982; 257(7): 3409–3413. pmid:7061488
View Article
PubMed/NCBI
Google Scholar

[34] View Article

[35] PubMed/NCBI

[36] Google Scholar

[ref10] 10. Pola A, Sadananthan SA, Yaligar J, Nagarajan V, Han WP, Kuchel PW, et al. Skeletal muscle lipid metabolism studied by advanced magnetic resonance spectroscopy. Prog Nucl Magn Reson Spectrosc. 2012; 65: 66–76. pmid:22781315
View Article
PubMed/NCBI
Google Scholar

[38] View Article

[39] PubMed/NCBI

[40] Google Scholar

[ref11] 11. Ludwig C, Viant MR. Two-dimensional J-resolved NMR spectroscopy: Review of a key methodology in the metabolomics toolbox. Phytochem Anal. 2010; 21(1): 22–32. pmid:19904730
View Article
PubMed/NCBI
Google Scholar

[42] View Article

[43] PubMed/NCBI

[44] Google Scholar

[ref12] 12. Terpstra M, Andersen PM, Gruetter R. Localized eddy current compensation using quantitative field mapping. J Magn Reson. 1998; 131(1): 139–143. pmid:9533916
View Article
PubMed/NCBI
Google Scholar

[46] View Article

[47] PubMed/NCBI

[48] Google Scholar

[ref13] 13. Kim DH, Adalsteinsson E, Glover GH, Spielman DM. Regularized higher-order in vivo shimming. Magn Reson Med. 2002; 48(4): 715–722. pmid:12353290
View Article
PubMed/NCBI
Google Scholar

[50] View Article

[51] PubMed/NCBI

[52] Google Scholar

[ref14] 14. Springer F, Machann J, Schwenzer NF, Ballweg V, Wurslin C, Schneider JH, et al. Quantitative assessment of intrahepatic lipids using fat-selective imaging with spectral-spatial excitation and in-/opposed-phase gradient echo imaging techniques within a study population of extremely obese patients feasibility on a short, wide-bore MR scanner. Invest Radiol. 2010; 45(8): 484–490. pmid:20479651
View Article
PubMed/NCBI
Google Scholar

[54] View Article

[55] PubMed/NCBI

[56] Google Scholar

[ref15] 15. Tugnoli V, Schenetti L, Mucci A, Nocetti L, Toraci C, Mavilla L, et al. A comparison between in vivo and ex vivo HR-MAS ¹H MR spectra of a pediatric posterior fossa lesion. Int J Mol Med. 2005; 16(2): 301–307. pmid:16012766
View Article
PubMed/NCBI
Google Scholar

[58] View Article

[59] PubMed/NCBI

[60] Google Scholar

[ref16] 16. Righi V, Durante C, Cocchi M, Calabrese C, Di Febo G, Lecce F, et al. Discrimination of healthy and neoplastic human colon tissues by ex vivo HR-MAS NMR spectroscopy and chemometric analyses. J Proteome Res. 2009; 8(4): 1859–1869. pmid:19714812
View Article
PubMed/NCBI
Google Scholar

[62] View Article

[63] PubMed/NCBI

[64] Google Scholar

[ref17] 17. Hamaed H, Pawlowski JM, Cooper BFT, Fu RQ, Eichhorn SH, Schurko RW. Application of solid-state ³⁵Cl NMR to the structural characterization of hydrochloride pharmaceuticals and their polymorphs. J Am Chem Soc. 2008; 130(33): 11056–11065. pmid:18656917
View Article
PubMed/NCBI
Google Scholar

[66] View Article

[67] PubMed/NCBI

[68] Google Scholar

[ref18] 18. Sitter B, Sonnewald U, Spraul M, Fjosne HE, Gribbestad IS. High-resolution magic angle spinning MRS of breast cancer tissue. Nmr Biomed. 2002; 15(5): 327–337. pmid:12203224
View Article
PubMed/NCBI
Google Scholar

[70] View Article

[71] PubMed/NCBI

[72] Google Scholar

[ref19] 19. Nestor G, Bankefors J, Schlechtriem C, Brannas E, Pickova J, Sandstrom C. High-resolution H-1 magic angle spinning NMR spectroscopy of intact arctic char (salvelinus alpinus) muscle. quantitative analysis of n-3 fatty acids, EPA and DHA. J Agr Food Chem. 2010; 58(20): 10799–10803.
View Article
Google Scholar

[74] View Article

[75] Google Scholar

[ref20] 20. Cai HH, Chen YS, Cui XH, Cai SH, Chen Z. High-Resolution H-1 NMR spectroscopy of fish muscle, eggs and small whole fish via hadamard-encoded intermolecular multiple-quantum coherence. Plos One. 2014; 9(1): e86422. pmid:24466083
View Article
PubMed/NCBI
Google Scholar

[77] View Article

[78] PubMed/NCBI

[79] Google Scholar

[ref21] 21. Vathyam S, Lee S, Warren WS. Homogeneous NMR spectra in inhomogeneous fields. Science. 1996; 272(5258): 92–96. pmid:8600541
View Article
PubMed/NCBI
Google Scholar

[81] View Article

[82] PubMed/NCBI

[83] Google Scholar

[ref22] 22. Lin YY, Ahn S, Murali N, Brey W, Bowers CR, Warren WS. High-resolution, > 1 GHz NMR in unstable magnetic fields. Phys Rev Lett. 2000; 85(17): 3732–3735. pmid:11030993
View Article
PubMed/NCBI
Google Scholar

[85] View Article

[86] PubMed/NCBI

[87] Google Scholar

[ref23] 23. Chen Z, Chen ZW, Zhong JH. High-resolution NMR spectra in inhomogeneous fields via IDEAL (intermolecular dipolar-interaction enhanced all lines) method. J Am Chem Soc. 2004; 126(2): 446–447. pmid:14719924
View Article
PubMed/NCBI
Google Scholar

[89] View Article

[90] PubMed/NCBI

[91] Google Scholar

[ref24] 24. Huang YQ, Cal SH, Zhang ZY, Chen Z. High-resolution two-dimensional J-resolved NMR spectroscopy for biological systems. Biophys J. 2014; 106(9): 2061–2070. pmid:24806938
View Article
PubMed/NCBI
Google Scholar

[93] View Article

[94] PubMed/NCBI

[95] Google Scholar

[ref25] 25. Bottomley PA. Spatial localization in NMR-spectroscopy in vivo. Ann Ny Acad Sci. 1987; 508: 333–348. pmid:3326459
View Article
PubMed/NCBI
Google Scholar

[97] View Article

[98] PubMed/NCBI

[99] Google Scholar

[ref26] 26. Liu ML, Mao XA, Ye CH, Huang H, Nicholson JK, Lindon JC. Improved WATERGATE pulse sequences for solvent suppression in NMR spectroscopy. J Magn Reson. 1998; 132(1): 125–129.
View Article
Google Scholar

[101] View Article

[102] Google Scholar

[ref27] 27. Zheng G, Price WS. Solvent signal suppression in NMR. Prog Nucl Magn Reson Spectrosc. 2010; 56(3): 267–288. pmid:20633355
View Article
PubMed/NCBI
Google Scholar

[104] View Article

[105] PubMed/NCBI

[106] Google Scholar

[ref28] 28. Ahn S, Warren WS, Lee S. Quantum treatment of intermolecular multiple-quantum coherences with intramolecular J coupling in solution NMR. J Magn Reson. 1997; 128(2): 114–129. pmid:9356265
View Article
PubMed/NCBI
Google Scholar

[108] View Article

[109] PubMed/NCBI

[110] Google Scholar

[ref29] 29. Lee S, Richter W, Vathyam S, Warren W. Quantum treatment of the effects of dipole–dipole interactions in liquid nuclear magnetic resonance. J Chem Phys. 1996; 105(3): 874–900.
View Article
Google Scholar

[112] View Article

[113] Google Scholar

[ref30] 30. Thomas MA, Ryner LN, Mehta MP, Turski PA, Sorenson JA. Localized 2D J-resolved ¹H MR spectroscopy of human brain tumors in vivo. J Magn Reson Imaging. 1996; 6(3): 453–459. pmid:8724410
View Article
PubMed/NCBI
Google Scholar

[115] View Article

[116] PubMed/NCBI

[117] Google Scholar

[ref31] 31. Ernst RR, Bodenhausen G, Wokaun A. Principles of nuclear magnetic resonance in one and two dimensions. Oxford: Clarendon Press; 1987.

[ref32] 32. Balla DZ, Melkus G, Faber C. Spatially localized intermolecular zero-quantum coherence spectroscopy for in vivo applications. Magn Reson Med. 2006; 56(4): 745–753. pmid:16897767
View Article
PubMed/NCBI
Google Scholar

[120] View Article

[121] PubMed/NCBI

[122] Google Scholar

[ref33] 33. Govindaraju V, Young K, Maudsley AA. Proton NMR chemical shifts and coupling constants for brain metabolites. Nmr Biomed. 2000; 13(3): 129–153. pmid:10861994
View Article
PubMed/NCBI
Google Scholar

[124] View Article

[125] PubMed/NCBI

[126] Google Scholar

[ref34] 34. Cui XH, Bao JF, Huang YQ, Cai SH, Chen Z. In vivo spatially localized high resolution ¹H MRS via intermolecular single-quantum coherence of rat brain at 7 T. J Magn Reson Imaging. 2013; 37(2): 359–364. pmid:23034817
View Article
PubMed/NCBI
Google Scholar

[128] View Article

[129] PubMed/NCBI

[130] Google Scholar

[ref35] 35. Faber C. Solvent-localized NMR spectroscopy using the distant dipolar field: A method for NMR separations with a single gradient. J Magn Reson. 2005; 176(1): 120–124. pmid:15990343
View Article
PubMed/NCBI
Google Scholar

[132] View Article

[133] PubMed/NCBI

[134] Google Scholar

[ref36] 36. Balla DZ, Faber C. Localized intermolecular zero-quantum coherence spectroscopy in vivo. Concept Magn Reson A. 2008; 32A(2): 117–133.
View Article
Google Scholar

[136] View Article

[137] Google Scholar

[ref37] 37. Gribbestad IS, Aursand M, Martinez I. High-resolution ¹H magnetic resonance spectroscopy of whole fish, fillets and extracts of farmed Atlantic salmon (Salmo salar) for quality assessment and compositional analyses. Aquaculture. 2005; 250(1–2): 445–457.
View Article
Google Scholar

[139] View Article

[140] Google Scholar

[ref38] 38. Sack M, Wetterling F, Sartorius A, Ende G, Weber-Fahr W. Signal-to-noise ratio of a mouse brain ¹³C CryoProbe^TM system in comparison with room temperature coils: Spectroscopic phantom and in vivo results. Nmr Biomed. 2014; 27(6): 709–715. pmid:24692120
View Article
PubMed/NCBI
Google Scholar

[142] View Article

[143] PubMed/NCBI

[144] Google Scholar

[ref39] 39. Hall EL, Stephenson MC, Price D, Morris PG. Methodology for improved detection of low concentration metabolites in MRS: Optimised combination of signals from multi-element coil arrays. Neuroimage. 2014; 86: 35–42. pmid:23639258
View Article
PubMed/NCBI
Google Scholar

[146] View Article

[147] PubMed/NCBI

[148] Google Scholar

[ref40] 40. Tal A, Frydman L. Single-scan multidimensional magnetic resonance. Prog Nucl Magn Reson Spectrosc. 2010; 57(3): 241–292. pmid:20667401
View Article
PubMed/NCBI
Google Scholar

[150] View Article

[151] PubMed/NCBI

[152] Google Scholar

[ref41] 41. Träber F, Block W, Freymann N, Gür O, Kucinski T, Hammen T, et al. A multicenter reproducibility study of single-voxel ¹H-MRS of the medial temporal lobe. Eur Radiol. 2006; 16(5): 1096–1103. pmid:16416279
View Article
PubMed/NCBI
Google Scholar

[154] View Article

[155] PubMed/NCBI

[156] Google Scholar

Figures

Abstract

Background and Purpose

Materials and Methods

Results

Conclusion

Introduction

Theory

Methods and Materials

Aqueous Solution

Pig Brain Tissue

A Whole Fish

Results and Discussion

Conclusion

Acknowledgments

Author Contributions

References