Spectral Editing Research Articles

The DEPTQ+ Experiment: Leveling the DEPT Signal Intensities and Clean Spectral Editing for Determining CHn Multiplicities.

We propose a new 13C DEPTQ+ NMR experiment, based on the improved DEPTQ experiment, which is designed to unequivocally identify all carbon multiplicities (Cq, CH, CH2, and CH3) in two experiments. Compared to this improved DEPTQ experiment, the DEPTQ+ is shorter and the different evolution delays are designed as spin echoes, which can be tuned to different 1JCH values; this is especially valuable when a large range of 1JCH coupling constants is to be expected. These modifications allow (i) a mutual leveling of the DEPT signal intensities, (ii) a reduction in J cross-talk in the Cq/CH spectrum, and (iii) more consistent and cleaner CH2/CH3 edited spectra. The new DEPTQ+ is expected to be attractive for fast 13C analysis of small-to medium sized molecules, especially in high-throughput laboratories. With concentrated samples and/or by exploiting the high sensitivity of cryogenically cooled 13C NMR probeheads, the efficacy of such investigations may be improved, as it is possible to unequivocally identify all carbon multiplicities, with only one scan, for each of the two independent DEPTQ+ experiments and without loss of quality.

Spectral editing of alanine, serine, and threonine in uniformly labeled proteins based on frequency-selective homonuclear recoupling in solid-state NMR.

Spectral editing is crucial to simplify the crowded solid-state NMR spectra of proteins. New techniques are introduced to edit 13C-13C correlations of uniformly labeled proteins under moderate magic-angle spinning (MAS), based on our recent frequency-selective homonuclear recoupling sequences [Zhang et al., J. Phys. Chem. Lett. 2020, 11, 8077-8083]. The signals of alanine, serine, or threonine residues are selected out by selective 13Cα-13Cβ double-quantum filtering (DQF). The 13Cα-13Cβ correlations of alanine residues are selectively established with efficiency up to ~ 1.8 times that by dipolar-assisted rotational resonance (DARR). The techniques are shown in 2D/3D NCCX experiments and applied to the uniformly 13C, 15N labeled Aquaporin Z (AqpZ) membrane protein, demonstrating their potential to simplify spectral analyses in biological solid-state NMR.

Prospective frequency and motion correction for edited 1H magnetic resonance spectroscopy

The major inhibitory neurotransmitter gamma-aminobutyric acid (GABA) and the dominant antioxidant glutathione (GSH) both play a crucial role in brain functioning and are involved in several neurodegenerative and psychiatric diseases. Magnetic resonance spectroscopy (MRS) is a unique way to measure these neurometabolites non-invasively, but the measurement is highly sensitive to head movements, and especially in specific patient groups, motion stabilization in MRS could be valuable. Conventional MRS is acquired at relatively short echo times (TE), however, for unambiguous detection of GABA and GSH, spectral editing techniques are typically used. These depend on longer TEs and use frequency selective spectral editing pulses to separate the low-intensity peaks of GABA and GSH from overlapping resonances, but results in further increased motion sensitivity. Low-intensity metabolite peaks are usually edited one-by-one, however, simultaneous editing of multiple metabolites can be achieved using a Hadamard scheme, resulting in a substantial reduction in scan time. To investigate and correct for motion sensitivity in both conventional short-TE MRS (PRESS) and edited MRS (HERMES), we implemented a navigator-based prospective motion correction strategy including reacquisition of corrupted data. PRESS and HERMES spectra were acquired without motion, with motion with correction (repeated twice), and with motion without correction. Results indicate that when sufficient retrospective outlier removal is used, no significant differences in concentration and spectral quality were observed between motion conditions, even without prospective correction. HERMES spectral editing data showed to be more sensitive to motion, as significant differences in metabolite estimates and variability of spectral quality measures were observed for tCr, GABA+ and GSH when only retrospective outlier removal was applied. When using both prospective and retrospective correction, spectral quality was improved to almost the level of the no-motion acquisition. No differences in metabolite ratios for GABA and GSH could be observed when using motion correction. In conclusion, edited MRS showed to be more prone to motion artifacts, and prospective motion correction can restore most of the spectral quality in both conventional and edited MRS.

Isolated-alkene-linked porous organic polymers (BIT-POPs): facile synthesis via ROMP and distinguishing overlapping signals in solid-state 13C NMR

The chemical structures of novel isolated-alkene-linked porous organic polymers (named BIT-POPs) were investigated through spectral editing techniques based on solid-state NMR.

IMG-21. PROSPECTIVE PREOPERATIVE DETERMINATION OF ISOCITRATE DEHYDROGENASE MUTATION IN GLIOMAS USING SPECTRAL EDITING MAGNETIC RESONANCE SPECTROSCOPY

BACKGROUNDGliomas are the most common malignant brain tumors in children and adults. A subset of these tumors harbour mutations in the enzyme isocitrate dehydrogenase (IDH) which produces the novel oncometabolite 2-hydroxyglutarate (2HG). In general, patients with an IDH mutant glioma have a longer survival—often necessitating more re-treatment sessions over the span of a patient’s life and surveillance monitoring for tumor recurrence. The need to non-invasively detect early evidence of tumor recurrence is therefore heightened in this unique subset of patients with extended survival. As magnetic resonance spectroscopy (MRS) has been demonstrated to measure biochemical components of intracranial tumors using MRI, we conducted a study in 58 pre-operative adult patients to determine if a diagnosis of IDH mutant glioma could be made confidently using imaging data.METHODSPatients underwent neuroimaging for diagnosis or preoperative planning on a 3 tesla MR scanner. A MEGA-PRESS spectral editing technique was employed. Imaging findings were directly compared to post-operative histopathologic diagnosis. RESUTLS: For all patients with gliomas from grade II to IV, detection of 2-HG with MEGA-PRESS sequence had a sensitivity between 48% and 81%, specificity between 60% and 100%, PPV between 53% and 100% and NPV between 77% and 85% depending on the CRLB threshold. Among the different metabolite ratios, a 2-HG/NAA ratio >0.034 had the highest sensitivity and specificity, 86% and 73% respectively.DISCUSSIONMagnetic resonance spectroscopy (MRS) is an underused advanced MR technique that deserves consideration in pediatric neuro-oncology given its utility in non-invasively detecting malignant gliomas.

GSH and GABA decreases in IDH1-mutated low-grade gliomas detected by HERMES spectral editing at 3 T in vivo

Isocitrate dehydrogenase 1 (IDH1) mutational status is an important prognostic biomarker in gliomas. γ-aminobutyric acid (GABA) and reduced glutathione (GSH) play an important role in energy production, which is related to tumor progression. Hadamard Encoding and Reconstruction of Mega-Edited Spectroscopy (HERMES) is able to detect GABA and GSH in healthy controls. This study aims to examine GABA and GSH alterations in IDH1-mutated low-grade gliomas using HERMES. We prospectively enrolled 14 suspected low-grade gliomas and 6 healthy control patients in this study, all cases underwent a 3 T MRI scan, including T1-weighted imaging and HERMES acquisition with a volume of interest 3 × 3 × 3 cm3. HERMES detects a “GABA+” signal that includes contributions from macromolecules and homocarnosine. GABA+ and GSH in tumor foci (group 1), contralateral cerebral regions (group 2) and healthy controls (group 3) were quantified using Gannet. The fitting errors and SNR of HERMES for GABA+ and GSH were analyzed; FWHM of the unsuppressed water signal was also recorded. The Wilcoxon signed-rank test was performed to test for differences between contralateral GABA+ and GSH levels, and differences in GABA+, GSH and fitting errors/SNR between the three groups were analyzed using analysis of variance (ANOVA). Eleven IDH1-mutant low-grade gliomas (5 Female and 6 Male, age 33–69) and 6 healthy subjects (2 Female and 4 Male, age 35–60) were finally enrolled this study. The mean water linewidth across all subjects was 9.67 ± 2.28 Hz. The Wilcoxon signed-rank test revealed that GABA+ and GSH were decreased significantly in glioma foci compared with contralateral regions, whereas no differences were seen between the left and right regions in healthy controls. ANOVA showed that GABA+ and GSH levels in tumor were lower than contralaterally and in healthy controls, while no differences were observed between the contralateral healthy tissue and healthy controls. No differences of fitting errors or SNR were found between tumors, contralateral regions or healthy controls. Our results suggest that HERMES is a reliable tool to simultaneously measure GABA and GSH alterations in low-grade gliomas with IDH1 mutations.

Solid-state NMR spectroscopy identifies three classes of lipids in Cryptococcus neoformans melanized cell walls and whole fungal cells

A primary virulence-associated trait of the opportunistic fungal pathogen Cryptococcus neoformans is the production of melanin pigments that are deposited into the cell wall and interfere with the host immune response. Previously, our solid-state NMR studies of isolated melanized cell walls (melanin "ghosts") revealed that the pigments are strongly associated with lipids, but their identities, origins, and potential roles were undetermined. Herein, we exploited spectral editing techniques to identify and quantify the lipid molecules associated with pigments in melanin ghosts. The lipid profiles were remarkably similar in whole C. neoformans cells, grown under either melanizing or nonmelanizing conditions; triglycerides (TGs), sterol esters (SEs), and polyisoprenoids (PPs) were the major constituents. Although no quantitative differences were found between melanized and nonmelanized cells, melanin ghosts were relatively enriched in SEs and PPs. In contrast to lipid structures reported during early stages of fungal growth in nutrient-rich media, variants found herein could be linked to nutrient stress, cell aging, and subsequent production of substances that promote chronic fungal infections. The fact that TGs and SEs are the typical cargo of lipid droplets suggests that these organelles could be connected to C. neoformans melanin synthesis. Moreover, the discovery of PPs is intriguing because dolichol is a well-established constituent of human neuromelanin. The presence of these lipid species even in nonmelanized cells suggests that they could be produced constitutively under stress conditions in anticipation of melanin synthesis. These findings demonstrate that C. neoformans lipids are more varied compositionally and functionally than previously recognized.

Spectral editing in 1 H magnetic resonance spectroscopy: Experts' consensus recommendations.

Spectral editing in in vivo 1 H-MRS provides an effective means to measure low-concentration metabolite signals that cannot be reliably measured by conventional MRS techniques due to signal overlap, for example, γ-aminobutyric acid, glutathione and D-2-hydroxyglutarate. Spectral editing strategies utilize known J-coupling relationships within the metabolite of interest to discriminate their resonances from overlying signals. This consensus recommendation paper provides a brief overview of commonly used homonuclear editing techniques and considerations for data acquisition, processing and quantification. Also, we have listed the experts' recommendations for minimum requirements to achieve adequate spectral editing and reliable quantification. These include selecting the right editing sequence, dealing with frequency drift, handling unwanted coedited resonances, spectral fitting of edited spectra, setting up multicenter clinical trials and recommending sequence parameters to be reported in publications.

Preoperative Determination of Isocitrate Dehydrogenase Mutation in Gliomas Using Spectral Editing MRS: A Prospective Study.

The edited magnetic resonance spectroscopy (MRS) technique has not yet been formally evaluated for the in vivo detection of 2-hydroxyglutarate (2-HG) in patients with gliomas of various grades. To evaluate the diagnostic accuracy of edited MRS in the preoperative identification of the isocitrate dehydrogenase (IDH) mutation status in patients with gliomas. Prospective. Fifty-eight subjects (31 glioblastomas, 27 grade II and III gliomas). Mescher-Garwood (MEGA)-PRESS and routine clinical brain tumor MR sequences were used at 3T. Data were analyzed using an advanced method for accurate, robust, and efficient spectral fitting (AMARES) from jMRUI software. The amplitudes of the 2-HG, N-acetyl-aspartate (NAA), choline (Cho), and creatine/phosphocreatine (Cr) resonances were calculated with their associated Cramer-Rao lower bound (CRLB). The IDH1 R132H mutation status was assessed by immunohistochemistry for all patients. Patients with grades II and III gliomas with negative immunohistochemistry underwent DNA sequencing to further interrogate IDH mutation status. The differences in 2-HG amplitudes, 2-HG/NAA, 2-HG/Cho, and 2-HG/Cr between IDH-mutant and IDH-wildtype gliomas were assessed using Mann-Whitney U-tests. Receiver operating characteristic curve analysis was performed to evaluate the diagnostic accuracy of each parameter. The 2-HG amplitudes, 2-HG/NAA, and 2-HG/Cho were higher for IDH-mutant gliomas than IDH-wildtype gliomas (P < 0.007). Using a CRLB threshold <30%, a 2-HG cutoff greater than 0 had a sensitivity of 80% (95% confidence interval [CI]: 52-96%) and a specificity of 81% (95% CI: 54-96%) in identifying IDH-mutant gliomas. In the subset of patients with grades II and III gliomas, the sensitivity was 80% (95% CI: 52-96%) and specificity was 100% (95% CI: 40-100%). Among 2-HG ratios, the highest AUC for the identification of IDH mutant status was achieved using the 2-HG/NAA (AUC = 0.8, 95% CI 0.67-.89). Preoperative edited MRS appears to be able to help identify IDH-mutant gliomas with high specificity. Level of Evidence 1 Technical Efficacy Stage 2 J. MAGN. RESON. IMAGING 2021;53:416-426.

Local and Interregional Neurochemical Associations Measured by Magnetic Resonance Spectroscopy for Studying Brain Functions and Psychiatric Disorders.

Magnetic resonance spectroscopy (MRS) studies have found significant correlations among neurometabolites (e.g., between glutamate and GABA) across individual subjects and altered correlations in neuropsychiatric disorders. In this article, we discuss neurochemical associations among several major neurometabolites which underpin these observations by MRS. We also illustrate the role of spectral editing in eliminating unwanted correlations caused by spectral overlapping. Finally, we describe the prospects of mapping macroscopic neurochemical associations across the brain and characterizing excitation–inhibition balance of neural networks using glutamate- and GABA-editing MRS imaging.

Determination of Brain Metabolite T1 Without Interference From Macromolecule Relaxation.

J-coupled metabolites are often measured at a predetermined echo time in the presence of macromolecule signals, which complicates the measurement of metabolite T1 . To evaluate the feasibility and benefits of measuring metabolite T1 relaxation times without changing the overlapping macromolecule baseline signals. Prospective. Five healthy volunteers (three females and two males; age = 27 ± 7 years). 7T scanner using a point resolved spectroscopy (PRESS)-based spectral editing MR spectroscopy (MRS) sequence with inversion recovery (IR). F-tests were performed to evaluate if the new approach, which fitted all the spectra together and used the same baselines for the three different IR settings, significantly reduced the variances of the metabolite T1 values compared to a conventional fitting approach. Cramer-Rao lower bound (CRLB), within-subject coefficient of variation, and F-test. The T1 relaxation times of N-acetylaspartate (NAA), total creatine (tCr), total choline (tCho), myo-inositol (mI), and glutamate (Glu) were determined with CRLB values below 6%. Glutamine (Gln) T1 was determined with a 17% CRLB, and the T1 of γ-aminobutyric acid (GABA) was determined with a 34% CRLB. The new approach significantly reduced the variances (F-test P < 0.05) of NAA, Glu, Gln, tCr, tCho, and mI T1 s compared to the conventional approach. Keeping macromolecule signals intact by using only long IR times allowed the use of a single macromolecule spectral model for different IR settings and significantly reduced the variances of NAA, Glu, Gln, tCr, tCho, and mI T1 s. 1 TECHNICAL EFFICACY STAGE: 1.

Ex vivo Comprehensive Multiphase NMR of whole organisms: A complementary tool to in vivo NMR

Nuclear Magnetic Resonance (NMR) spectroscopy is a non-invasive analytical technique which allows for the study of intact samples. Comprehensive Multiphase NMR (CMP-NMR) combines techniques and hardware from solution state and solid state NMR to allow for the holistic analysis of all phases (i.e. solutions, gels and solids) in unaltered samples. This study is the first to apply CMP-NMR to deceased, intact organisms and uses 13C enriched Daphnia magna (water fleas) as an example. D. magna are commonly used model organisms for environmental toxicology studies. As primary consumers, they are responsible for the transfer of nutrients across trophic levels, and a decline in their population can potentially impact the entire freshwater aquatic ecosystem. Though in vivo research is the ultimate tool to understand an organism’s most biologically relevant state, studies are limited by conditions (i.e. oxygen requirements, limited experiment time and reduced spinning speed) required to keep the organisms alive, which can negatively impact the quality of the data collected. In comparison, ex vivo CMP-NMR is beneficial in that; organisms do not need oxygen (eliminating air holes in rotor caps and subsequent evaporation); samples can be spun faster, leading to improved spectral resolution; more biomass per sample can be analyzed; and experiments can be run for longer. In turn, higher quality ex vivo NMR, can provide more comprehensive NMR assignments, which in many cases could be transferred to better understand less resolved in vivo signals. This manuscript is divided into three sections: 1) multiphase spectral editing techniques, 2) detailed metabolic assignments of 2D NMR of 13C enriched D. magna and 3) multiphase biological changes over different life stages, ages and generations of D. magna. In summary, ex vivo CMP-NMR proves to be a very powerful approach to study whole organisms in a comprehensive manner and should provide very complementary information to in vivo based research.

Osprey: Open-source processing, reconstruction & estimation of magnetic resonance spectroscopy data

BackgroundProcessing and quantitative analysis of magnetic resonance spectroscopy (MRS) data are far from standardized and require interfacing with third-party software. Here, we present Osprey, a fully integrated open-source data analysis pipeline for MRS data, with seamless integration of pre-processing, linear-combination modelling, quantification, and data visualization. New MethodOsprey loads multiple common MRS data formats, performs phased-array coil combination, frequency-and phase-correction of individual transients, signal averaging and Fourier transformation. Linear combination modelling of the processed spectrum is carried out using simulated basis sets and a spline baseline. The MRS voxel is coregistered to an anatomical image, which is segmented for tissue correction and quantification is performed based upon modelling parameters and tissue segmentation. The results of each analysis step are visualized in the Osprey GUI. The analysis pipeline is demonstrated in 12 PRESS, 11 MEGA-PRESS, and 8 HERMES datasets acquired in healthy subjects. ResultsOsprey successfully loads, processes, models, and quantifies MRS data acquired with a variety of conventional and spectral editing techniques. Comparison with Existing Method(s)Osprey is the first MRS software to combine uniform pre-processing, linear-combination modelling, tissue correction and quantification into a coherent ecosystem. Compared to existing compiled, often closed-source modelling software, Osprey’s open-source code philosophy allows researchers to integrate state-of-the-art data processing and modelling routines, and potentially converge towards standardization of analysis. ConclusionsOsprey combines robust, peer-reviewed data processing methods into a modular workflow that is easily augmented by community developers, allowing the rapid implementation of new methods.

Signal enhancement of glutamine and glutathione by single-step spectral editing

A single-step spectral editing approach using an always-on editing pulse was proposed to enhance the signals of strongly coupled spins. Specifically, a single-step spectral editing sequence with an always-on editing pulse applied at 2.12 ppm was used to enhance glutamine (Gln) and glutathione (GSH) signals at TE = 56 ms on a 7 T scanner. Density matrix simulations demonstrated that the current method (TE = 56 ms) led to large signal enhancement of at least 61% for Gln and 51% for GSH compared to a previous single-step method (TE = 106 ms). Monte Carlo simulations showed that the current method reduced noise-originated variations by 31% for Gln and 26% for GSH compared to a previous three-step spectral editing method from which the present method was derived.

Quantitative measurement of redox state in human brain by 31 P MRS at 7T with spectral simplification and inclusion of multiple nucleotide sugar components in data analysis.

To develop a simplified method for quantitative measurement of NAD+ /NADH (nicotinamide adenine dinucleotides) levels in human brain by 31 P MRS without interference from the α-ATP signal and with inclusion of multiple UDP-sugar components. Simple pulse-acquire 31 P MR spectra were collected at 7T with and without a frequency-selective inversion pulse to remove the dominant α-ATP signal from the underlying NAD(H) signal. Careful inspection of the 31 P signal at -9.8 ppm previously assigned to UDP-glucose revealed multiple UDP-sugar components that must also be considered when deconvoluting the NAD(H) signal to quantify NAD+ and NADH. Finally, the overlapping NAD(H) and UDP(G) resonances were deconvoluted into individual components using Voigt lineshape analysis and UDP(G) modeling. The inversion-based spectral editing method enabled clean separation of the NAD(H) signal from the otherwise dominant α-ATP signal. In addition, the upfield signal near -9.8 ppm appears more "quartet-like" than a simple doublet consistent with contributions from other nucleotide sugars such as UDP-galactose, UDP-N-acetyl-galactosamine, and UDP-N-acetyl-glucosamine in addition to UDP-glucose. Deconvolution of the combined NAD(H) and UDP(G) signals showed that the measured NAD+ /NAD ratio was heavily influenced by UDP(G) modeling (7.5 ± 1.8 when the UDP(G) signal was fitted as multiple doublets versus 5.3 ± 0.6 when a simplified pseudo doublet model was used). In a test/re-test experiments separated by 2 weeks, consistent NAD+ /NADH ratios were measured in the brain of seven human subjects. The NAD+ /NADH ratio in human brain can be measured using 31 P MR spectra simplified by spectral editing and with inclusion of multiple UDP-sugar components in the data analysis.

Motion correction in magnetic resonance spectroscopy.

In vivo proton magnetic resonance spectroscopy and spectroscopic imaging (MRS/MRSI) are valuable tools to study normal and abnormal human brain physiology. However, they are sensitive to motion, due to strong crusher gradients, long acquisition times, reliance on high magnetic field homogeneity, and particular acquisition methods such as spectral editing. The effects of motion include incorrect spatial localization, phase fluctuations, incoherent averaging, line broadening, and ultimately quantitation errors. Several retrospective methods have been proposed to correct motion-related artifacts. Recent advances in hardware also allow prospective (real-time) correction of the effects of motion, including adjusting voxel location, center frequency, and magnetic field homogeneity. This article reviews prospective and retrospective methods available in the literature and their implications for clinical MRS/MRSI. In combination, these methods can attenuate or eliminate most motion-related artifacts and facilitate the acquisition of high-quality data in the clinical research setting.

CoHEARence of audible shapes\u2014a qualitative user study for coherent visual audio design with resynthesized shapes

One way to achieve an intuitive sound design are visual approaches for synthesis and sound collages. Therefore, during spectral synthesis and editing, the sound is designed in a visualization of the frequency domain. In order to create a coherent workflow between visuals and the resulting audio, the stimuli should be matched to each other. In this work, a qualitative user study is presented, which is supposed to show the intuitive understanding from the shape to the sound. The shape is hereby the spectral envelope. The general aim is to find out whether there is a connection between the visual shape and the subsequent auditory impression.

Formation of Char-Like, Fused-Ring Aromatic Structures from a Nonpyrogenic Pathway during Decomposition of Wheat Straw.

Fused-ring aromatics, important skeletal components of black carbon (BC), contribute to long-term carbon (C) sequestration in nature. They have previously been thought to be primarily formed by incomplete combustion of organic materials, whereas the nonpyrogenic origins are negligible. Using advanced solid-state 13C nuclear magnetic resonance (NMR), including recoupled long-range C-H dipolar dephasing, exchange with protonated and nonprotonated spectral editing (EXPANSE), and dipolar-dephased double-quantum/single-quantum (DQ/SQ) spectroscopy, we for the first time identify fused-ring aromatics that formed during the decomposition of wheat (Triticum sp.) straw in soil under aerobic, but not anaerobic conditions. The observed formation of polyaromatic units as plant litter decomposes provides direct evidence for humification. Moreover, the estimation of the annual flux of such nonpyrogenic BC could be equivalent to 3-12% of pyrogenic BC added to soils from all other sources. Our findings significantly extend the understanding of potential sources of fused-ring aromatic C and BC in soils as well as the global C cycle.

Multinuclear solid-state NMR of complex nitrogen-rich polymeric microcapsules: Weight fractions, spectral editing, component mixing, and persistent radicals

The molecular structure of a crosslinked nitrogen-rich resin made from melamine, urea, and aldehydes, and of microcapsules made from the reactive resin with multiple polymeric components in aqueous dispersion, has been analyzed by 13C, 13C{1H}, 1H–13C, 1H, 13C{14N}, and 15N solid-state NMR without isotopic enrichment. Quantitative 13C NMR spectra of the microcapsules and three precursor materials enable determination of the fractions of different components. Spectral editing of non-protonated carbons by recoupled dipolar dephasing, of CH by dipolar DEPT, and of C–N by 13C{14N} SPIDER resolves peak overlap and helps with peak assignment. It reveals that the N- and O-rich resin “imitates” the spectrum of polysaccharides such as chitin, cellulose, or Ambergum to an astonishing degree. 15N NMR can distinguish melamine from urea and guanazole, NC=O from COO, and primary from secondary amines. Such a comprehensive and quantitative analysis enables prediction of the elemental composition of the resin, to be compared with combustion analysis for validation. It also provides a reliable reference for iterative simulations of 13C NMR spectra from structural models. The conversion from quantitative NMR peak areas of structural components to the weight fractions of interest in industrial practice is derived and demonstrated. Upon microcapsule formation, 15N and 13C NMR consistently show loss of urea and aldehyde and an increase in primary amines while melamine is retained. NMR also made unexpected findings, such as imbedded crystallites in one of the resins, as well as persistent radicals in the microcapsules. The crystallites produce distinct sharp lines and are distinguished from liquid-like components by their strong dipolar couplings, resulting in fast dipolar dephasing. Fast 1H spin–lattice relaxation on the 35-ms time scale and characteristically non-exponential 13C spin–lattice relaxation indicate persistent radicals, confirmed by EPR. Through 1H spin diffusion, the mixing of components on the 5-nm scale was documented.

A molecular fluorophore in citric acid/ethylenediamine carbon dots identified and quantified by multinuclear solid-state nuclear magnetic resonance.

The composition of fluorescent polymer nanoparticles, commonly referred to as carbon dots, synthesized by microwave-assisted reaction of citric acid and ethylenediamine was investigated by 13 C, 13 C{1 H}, 1 H─13 C, 13 C{14 N}, and 15 N solid-state nuclear magnetic resonance (NMR) experiments. 13 C NMR with spectral editing provided no evidence for significant condensed aromatic or diamondoid carbon phases. 15 N NMR showed that the nanoparticle matrix has been polymerized by amide and some imide formation. Five small, resolved 13 C NMR peaks, including an unusual ═CH signal at 84 ppm (1 H chemical shift of 5.8 ppm) and ═CN2 at 155 ppm, and two distinctive 15 N NMR resonances near 80 and 160 ppm proved the presence of 5-oxo-1,2,3,5-tetrahydroimidazo[1,2-a]pyridine-7-carboxylic acid (IPCA) or its derivatives. This molecular fluorophore with conjugated double bonds, formed by a double cyclization reaction of citric acid and ethylenediamine as first shown by Y. Song, B. Yang, and coworkers in 2015, accounts for the fluorescence of the carbon dots. Cross-peaks in a 1 H─13 C HETCOR spectrum with brief 1 H spin diffusion proved that IPCA is finely dispersed in the polyamide matrix. From quantitative 13 C and 15 N NMR spectra, a high concentration (18 ± 2 wt%) of IPCA in the carbon dots was determined. A pronounced gradient in 13 C chemical-shift perturbations and peak widths, with the broadest lines near the COO group of IPCA, indicated at least partial transformation of the carboxylic acid of IPCA by amide or ester formation.