Deuteration Level Research Articles

DGet! An open source deuteration calculator for mass spectrometry data

DGet! is an open-source analysis package written in Python for calculating the degree of deuterium enrichment in isotopically labelled molecules using mass spectrometric data. The nuclear properties of deuterium make it a valuable tracer in metabolic studies and an excellent contrast agent in nuclear spectroscopies. Determination of molecular deuteration levels is typically performed using mass spectrometry, however software options to perform these calculations are scarce. The in-house scripts and spreadsheets currently used rarely account for isotopic interferences from 13C or multi-isotopic elements that impact deuteration calculations. DGet! removes isotopic interferences using de-convolution and both the isotopological makeup and overall deuteration level can be accurately recovered. The software is available as command line and web applications that take a molecular formula and mass spectrometry data and output a graphical representation of the degree of deuteration as well as the distribution of partially deuterated analogues. These applications are designed to be easy to use and enable superior characterisation of deuterated molecules for users of all levels of expertise, without the limitations of techniques currently used by the majority of deuteration laboratories and researchers.Graphical

Untargeted Lipidomics after D2O Administration Reveals the Turnover Rate of Individual Lipids in Various Organs of Living Organisms.

The administration of low doses of D2O to living organisms was used for decades for the investigation of metabolic pathways and for the measurement of the turnover rate for specific compounds. Usually, the investigation of the deuterium uptake in lipids is performed by measuring the deuteration level of the palmitic acid residue using GC-MS instruments, and to our knowledge, the application of the modern untargeted LC-MS/MS lipidomics approaches was only reported a few times. Here, we investigated the deuterium uptake for >500 lipids for 13 organs and body liquids of mice (brain, lung, heart, liver, kidney, spleen, plasma, urine, etc.) after 4 days of 100% D2O administration. The maximum deuteration level was observed in the liver, plasma, and lung, while in the brain and heart, the deuteration level was lower. Using MS/MS, we demonstrated the incorporation of deuterium in palmitic and stearic fragments in lipids (PC, PE, TAG, PG, etc.) but not in the corresponding free forms. Our results were analyzed based on the metabolic pathways of lipids.

Catalytic Vilsmeier-Haack Reactions for C1-Deuterated Formylation of Indoles.

The Vilsmeier-Haack reaction is a powerful tool to introduce formyl groups into electron-rich arenes, but its wide application is significantly restricted by stoichiometric employment of caustic POCl3. Herein, we reported a catalytic version of the Vilsmeier-Haack reaction enabled by a P(III)/P(V)═O cycle. This catalytic reaction provides a facile and efficient route for the direct construction of C1-deuterated indol-3-carboxaldehyde under mild conditions with stoichiometric DMF-d7 as the deuterium source. The products feature a remarkably higher deuteration level (>99%) than previously reported ones and are not contaminated by the likely unselective deuteration at other sites. The present transformation can also be used to transfer other carbonyl groups. Further downstream derivatizations of these deuterated products manifested their potential applications in the synthesis of deuterated bioactive molecules. Mechanistic insight was disclosed from studies of kinetics and intermediates.

Deuteration for biological SANS: Case studies, success and challenges in chemistry and biology.

Small angle neutron scattering is a powerful complementary technique in structural biology. It generally requires, or benefits from, deuteration to achieve its unique potentials. Molecular deuteration has become a mature expertise, with deuteration facilities located worldwide to support access to the technique for a wide breadth of structural biology and life sciences. The sorts of problems well answered by small angle scattering and deuteration involve large (>10Å) scale flexible movements, and this approach is best used where high-resolution methods (crystallography, NMR, cryo-EM) leave questions unanswered. This chapter introduces deuteration, reviewing biological deuteration of proteins, lipids and sterols, and then steps through the ever-expanding range of deuterated molecules being produced by chemical synthesis and enabling sophisticated experiments using physiologically relevant lipids. Case studies of recent successful use of deuteration may provide illustrative examples for strategies for future experiments. We discuss issues of nomenclature for synthesised molecules of novel labeling and make recommendations for their naming. We reflect on our experiences, with cost associated with achieving an arbitrary deuteration level, and on the benefits of experimental co-design by user scientist, deuteration scientist, and neutron scattering scientist working together. Although methods for biological and chemical deuteration are published in the public domain, we recommend that the best method to deuterate is to engage with a deuteration facility.

Diffusion equation for the longitudinal spectral diffusion: the case of the RIDME experiment.

Relaxation-induced dipolar modulation enhancement (RIDME) time trace shapes reveal linear scaling with the proton concentration in homogeneous glassy samples. We describe here an approximate diffusion equation-based analysis of such data, which uses only two fit parameters and allows for global data fitting with good accuracy. By construction, the approach should be transferable to other pulse EPR experiments with longitudinal mixing block(s) present. The two fit parameters appear to be sensitive to the type of the glassy matrix and can be thus used for sample characterisation. The estimates suggest that the presented technique should be sensitive to protons at distances up to 3 nm from the electron spin at a 90% matrix deuteration level. We propose that a structural method might be developed based on such an intermolecular hyperfine (ih-)RIDME technique, which would be useful, for instance, in structural biology or dynamic nuclear polarisation experiments.

Deuterated Polycyclic Aromatic Hydrocarbons in the Interstellar Medium: The C–D Band Strengths of Multideuterated Species

Abstract Observationally, the interstellar gas-phase abundance of deuterium (D) is considerably depleted and the missing D atoms are often postulated to have been locked up into carbonaceous solids and polycyclic aromatic hydrocarbon (PAH) molecules. An accurate knowledge of the fractional amount of D (relative to H) tied up in carbon dust and PAHs has important cosmological implications since D originated exclusively from the Big Bang and the present-day D abundance, after accounting for the astration it has experienced during Galactic evolution, provides essential clues to the primordial nucleosynthesis and the cosmological parameters. To quantitatively explore the extent to which PAHs could possibly accommodate the observed D depletion, we have previously quantum-chemically computed the infrared vibrational spectra of monodeuterated PAHs and derived the mean intrinsic band strengths of the 3.3 μm C–H stretch (A 3.3) and the 4.4 μm C–D stretch (A 4.4). Here we extend our previous work to multideuterated PAH species of different deuterations, sizes, and structures. We find that both the intrinsic band strengths A 3.3 and A 4.4 (on a per-unit-bond basis) and their ratios A 4.4/A 3.3 not only show little variation among PAHs of different deuterations, sizes, and structures, they are also closely similar to that of monodeuterated PAHs. Therefore, a PAH deuteration level (i.e., the fraction of peripheral atoms attached to C atoms in the form of D) of ∼2.4% previously estimated from the observed 4.4 to 3.3 μm band ratio based on the A 4.4/A 3.3 ratio of monodeuterated PAHs is robust.

Capillary Electrophoresis-based Hydrogen/Deuterium Exchange for Conformational Characterization of Proteins with Top-down Mass Spectrometry.

Resolving conformational heterogeneity of multiple protein states that coexist in solution remains one of the main obstacles in the characterization of protein therapeutics and the determination of the conformational transition pathways critical for biological functions, ranging from molecular recognition to enzymatic catalysis. Hydrogen/deuterium exchange (HDX) reaction coupled with top-down mass spectrometric (MS) analysis provides a means to characterize protein higher-order structures and dynamics in a conformer-specific manner. The conformational resolving power of this technique is highly dependent on the efficiencies of separating protein states at the intact protein level and minimizing the residual non-deuterated protic content during the HDX reactions. Here we describe a capillary electrophoresis (CE)-based variant of the HDX MS approach that aims to improve the conformational resolution. In this approach, proteins undergo HDX reactions while migrating through a deuterated background electrolyte solution (BGE) during the capillary electrophoretic separation. Different protein states or proteoforms that coexist in solution can be efficiently separated based on their differing charge-to-size ratios. The difference in electrophoretic mobility between proteins and protic solvent molecules minimizes the residual non-deuterated solvent, resulting in a nearly complete deuterating environment during the HDX process. The flow-through microvial CE-MS interface allows efficient electrospray ionization of the eluted protein species following a rapid mixing with the quenching and denaturing modifier solution at the outlet of the sprayer. The online top-down MS analysis measures the global deuteration level of the eluted intact protein species, and subsequently, the deuteration of their gas-phase fragments. This paper demonstrates this approach in differential HDX for systems, including the natural protein variants coexisting in milk.

Numerical analysis of the DKDP-based high-energy optical parametric chirped pulse amplifier for a 100 PW class laser.

An optical parametric chirped pulse amplifier (OPCPA) based on a large-aperture DKDP crystal and pumped by a 10 kJ level Nd:glass laser can serve as the final amplifier for a 100 PW level laser. A comprehensive numerical investigation on such a high-energy OPCPA is presented in this work. The effects on the efficiency-bandwidth product induced by the deuteration level, absorption loss, temperature variation, and optimization of zero-phase-mismatch wavelength (ZPMW) are analyzed in detail. Based on the analysis above, a three-dimensional numerical simulation taking into account the effects of pumping depletion, diffraction, and walk-off shows that, by optimization of ZPMW, broadband (over 210 nm spectral width in FWHM) and high efficiency (${\gt}37\%$) amplification can be realized in the DKDP crystal even with a moderate deuteration level of 70%, which can relax the requirement of a high deuteration level in a large-aperture DKDP crystal. The numerical analysis can provide meaningful guidance for the design and construction of 100 PW class laser systems.

Characterization of partially deuterated KDP crystals using two-wavelength phase-matching angles

Partially deuterated potassium dihydrogen phosphate (DKDP) allows for the optical parametric chirped-pulse amplification of high-energy broadband optical pulses to generate ultrashort, high-peak-power pulses. Modeling and experimental optimization of the noncollinear interaction geometry require the combination of a model for the deuteration-dependent optical indices and knowledge of the deuteration level of the DKDP crystal being used. We study and demonstrate a novel experimental technique that determines the deuteration level of a DKDP crystal consistent with a specific index model, based on the phase-matching angles measured at two wavelengths. Simulations and experiments show that the simple two-wavelength technique allows for consistent results with three different index models, and in particular, for the characterization of four crystals with deuteration levels ranging from 70% to 98%.

First detection of NHD and ND2 in the interstellar medium

Context. Deuterium fractionation processes in the interstellar medium (ISM) have been shown to be highly efficient in the family of nitrogen hydrides. To date, observations have been limited to ammonia (NH2D, NHD2, ND3) and imidogen radical (ND) isotopologues. Aims. We want to explore the high-frequency windows offered by the Herschel Space Observatory to search for deuterated forms of the amidogen radical NH2 and to compare the observations against the predictions of our comprehensive gas-grain chemical model. Methods. Making use of the new molecular spectroscopy data recently obtained at high frequencies for NHD and ND2, we searched for both isotopologues in the spectral survey toward the Class 0 protostar IRAS 16293-2422, a source in which NH3, NH, and their deuterated variants have previously been detected. We used the observations carried out with HIFI (Heterodyne Instrument for the Far-Infrared) in the framework of the key program “Chemical Herschel surveys of star forming regions” (CHESS). Results. We report the first detection of interstellar NHD and ND2. Both species are observed in absorption against the continuum of the protostar. From the analysis of their hyperfine structure, accurate excitation temperature and column density values are determined. The latter were combined with the column density of the parent species NH2 to derive the deuterium fractionation in amidogen. We find a high deuteration level of amidogen radical in IRAS 16293-2422, with a deuterium enhancement about one order of magnitude higher than that predicted by earlier astrochemical models. Such a high enhancement can only be reproduced by a gas-grain chemical model if the pre-stellar phase preceding the formation of the protostellar system has a long duration: on the order of one million years. Conclusions. The amidogen D/H ratio measured in the low-mass protostar IRAS 16293-2422 is comparable to that derived for the related species imidogen and much higher than that observed for ammonia. Additional observations of these species will provide more insights into the mechanism of ammonia formation and deuteration in the ISM. Finally, we indicate the current possibilities to further explore these species at submillimeter wavelengths.

Novel approach of qNMR workflow by standardization using 2H integral: Application to any intrinsic calibration standard

Quantitative nuclear magnetic resonance (qNMR) is routinely performed by the internal or external standardization. The manuscript describes a simple alternative to these common workflows by using NMR signal of another active nuclei of calibration compound. For example, for any arbitrary compound quantification by NMR can be based on the use of an indirect concentration referencing that relies on a solvent having both 1H and 2H signals. To perform high-quality quantification, the deuteration level of the utilized deuterated solvent has to be estimated.In this contribution the new method was applied to the determination of deuteration levels in different deuterated solvents (MeOD, ACN, CDCl3, acetone, benzene, DMSO-d6). Isopropanol-d6, which contains a defined number of deuterons and protons, was used for standardization. Validation characteristics (precision, accuracy, robustness) were calculated and the results showed that the method can be used in routine practice. Uncertainty budget was also evaluated. In general, this novel approach, using standardization by 2H integral, benefits from reduced sample preparation steps and uncertainties, and can be applied in different application areas (purity determination, forensics, pharmaceutical analysis, etc.).

Proton-deuterium exchange of acetone catalyzed in imidazolium-based ionic liquid-D2O mixtures.

The reaction of the proton–deuterium exchange of acetone in imidazolium-based ionic liquid (IL)–deuterium oxide mixtures was studied in detail via NMR spectroscopy. Certain ILs exhibit considerable catalytic properties and contribute to the course of reaction up to the complete deuteration. The efficiency of deuterium exchange crucially depends on the features of ILs; the type of anion and chain length of cation. The linear secondary isotope effects on the NMR chemical shifts of the 13C atoms in acetone were observed depending on the deuteration level of the molecule.

High-sensitivity maps of molecular ions in L1544

Context. The deuterium fraction in low-mass prestellar cores is a good diagnostic indicator of the initial phases of star formation, and is also a fundamental quantity to infer the ionisation degree in these objects. Aims. With the analysis of multiple transitions of N2H+, N2D+, HC18O+, and DCO+ we are able to determine the molecular column density maps and the deuterium fraction in N2H+ and HCO+ toward the prototypical prestellar core L1544. This is the preliminary step to derive the ionisation degree in the source. Methods. We used a non-local thermodynamic equilibrium (non-LTE) radiative transfer code combined with the molecular abundances derived from a chemical model to infer the excitation conditions of all the observed transitions. This allowed us to derive reliable maps of the column density of each molecule. The ratio between the column density of a deuterated species and its non-deuterated counterpart gives the sought-after deuteration level. Results. The non-LTE analysis confirms that, for the molecules analysed, higher-J transitions are characterised by excitation temperatures that are ≈1–2 K lower than those of the lower-J transitions. The chemical model that provides the best fit to the observational data predicts the depletion of N2H+ and to a lesser extent of N2D+ in the innermost region. The peak values for the deuterium fraction that we find are D/HN2H+ = 0.26−0.14+0.15 and D/HHCO+=0.035−0.012+0.015, in good agreement with previous estimates in the source.

3D Residual Strain in DKDP Crystals by Neutron Diffraction

AbstractNeutron diffraction technique is applied to study the three‐dimensional (3D) residual strains in K(H1−xDx)2PO4 (DKDP) crystals with various deuterium contents (0 ≤ x ≤ 0.99). It is found that the residual strains in the crystalline coordinate are at the magnitude of 10−3 to 10−4. The results show that the level of the macro‐strain in the crystal bulk is independent of the deuteration level. However, when the deuteration level is below 50%, the average of micro‐strain increases first with the increasing of deuteration level, then decreases when the deuteration level is in excess of 50%. According to the residual strains, the internal stresses are calculated through σij = Cijklεkl. Finally, the potential sources of the stresses in crystals are discussed and analyzed.

Threshold protective effect of deuterated polyunsaturated fatty acids on peroxidation of lipid bilayers.

Autoxidation of polyunsaturated fatty acids (PUFAs) damages lipid membranes and generates numerous toxic by-products implicated in neurodegeneration, aging, and other pathologies. Abstraction of bis-allylic hydrogen atoms is the rate-limiting step of PUFA autoxidation, which is inhibited by replacing bis-allylic hydrogens with deuterium atoms (D-PUFAs). In cells, the presence of a relatively small fraction of D-PUFAs among natural PUFAs is sufficient to effectively inhibit lipid peroxidation (LPO). Here, we investigate the effect of various D-PUFAs on the stability of liposomes under oxidative stress conditions. The permeability of vesicle membranes to fluorescent dyes was measured as a proxy for bilayer integrity, and the formation of conjugated dienes was monitored as a proxy for LPO. Remarkably, both approaches reveal a similar threshold for the protective effect of D-PUFAs in liposomes. We show that protection rendered by D-PUFAs depends on the structure of the deuterated fatty acid. Our findings suggest that protection of PUFAs against autoxidation depends on the total level of deuterated bi-sallylic (CD2 ) groups present in the lipid bilayer. However, the phospholipid containing 6,6,9,9,12,12,15,15,18,18-d10 -docosahexaenoic acid exerts a stronger protective effect than should be expected from its deuteration level. These findings further support the application of D-PUFAs as preventive/therapeutic agents in numerous pathologies that involve LPO.

POPPeT: a New Method to Predict the Protection Factor of Backbone Amide Hydrogens

Hydrogen exchange (HX) has become an important tool to monitor protein structure and dynamics. The interpretation of HX data with respect to protein structure requires understanding of the factors that influence exchange. Simulated protein structures can be validated by comparing experimental deuteration profiles with the profiles derived from the modeled protein structure. To do this, we propose here a new method, POPPeT, for protection factor prediction based on protein motions that enable HX. By comparing POPPeT with two existing methods, the phenomenological approximation and COREX, we show enhanced predictability measured at both protection factor and deuteration level. This method can be subsequently used by modeling strategies for protein structure prediction.Graphical ᅟ

A Study of the c-C3HD/c-C3H2 Ratio in Low-mass Star-forming Regions*

Abstract We use the deuteration of c-C3H2 to probe the physical parameters of starless and protostellar cores, related to their evolutionary states, and compare it to the N2H+-deuteration in order to study possible differences between the deuteration of C- and N-bearing species. We observed the main species c-C3H2, the singly and doubly deuterated species c-C3HD and c-C3D2, as well as the isotopologue c-H13CC2H toward 10 starless cores and five protostars in the Taurus and Perseus complexes. We examined the correlation between the N(c-C3HD)/N(c-C3H2) ratio and the dust temperature along with the H2 column density and the CO depletion factor. The resulting N(c-C3HD)/N(c-C3H2) ratio is, within error bars, consistent with 10% in all starless cores with detected c-C3HD. This also accounts for the protostars except for the source HH211, where we measure a high deuteration level of 23%. The deuteration of N2H+ follows the same trend but is considerably higher in the dynamically evolved core L1544. We find no significant correlation between the deuteration of c-C3H2 and the CO depletion factor among the starless and protostellar cores. Toward the latter the coolest objects show the largest deuterium fraction in c-C3H2. We show that the deuteration of c-C3H2 can trace the early phases of star formation and is comparable to that of N2H+. However, the largest c-C3H2 deuteration level is found toward protostellar cores, suggesting that while c-C3H2 is mainly frozen onto dust grains in the central regions of starless cores, active deuteration is taking place on ice.

The performance studies of DKDP crystals grown by a rapid horizontal growth method

A deuterated potassium dihydrogen phosphate (DKDP) crystal with about 70% deuterium level was grown by a rapid horizontal growth method with independent design equipment, which includes a continuous filtration system. The cooling program during crystal growth was designed according to a self-developed software to catch the size of growing crystal in real time. The crystal structure, optical performance and laser induced damage threshold (LIDT) of this DKDP crystal were investigated in this paper. The deuterium concentration of the crystal was confirmed by the neutron diffraction technique, which was effective and available in determining a complete range of deuteration level. The dielectric property was measured to evaluate the perfection of the lattice. The transmittance and LIDT were carried out further to evaluate the optical and functional properties of this DKDP crystal grown in the rapid horizontal growth technique. All of the detailed characterization for DKDP figured out that the 70% deuterated KDP crystal grown in this way had relatively good qualities.

Spectral gain investigation of large size OPCPA based on partially deuterated KDP

The Optical Parametric Chirped Pulse Amplification is one of the most promising techniques to deliver 20PW laser system. The already available KD*P in large size is a good candidate as nonlinear crystal. In this article we report the experimental analysis of the spectral small signal gain for KD*P at 70% deuteration level for different phase matching and non-collinear angle. The data is also compared with a theoretical model.

Deuterium Fractionation upon the Formation of Hexamethylenetetramines through Photochemical Reactions of Interstellar Ice Analogs Containing Deuterated Methanol Isotopologues

Abstract Hexamethylenetetramine (HMT) is a representative product after the photolysis of interstellar ice analogs containing methanol followed by warming-up to room temperature. Since interstellar methanol is often significantly enriched in deuterium (D), the HMT photoproduct is expected to inherit D atoms from deuterated methanol. However, D fractionation upon the formation of HMT is not well understood, especially when it is produced from partly deuterated methanol isotopologues such as CH2DOH and CH3OD. Here, we experimentally studied the composition of deuterated HMT (d n -HMT, where n is the number of D atoms) at the isotopologue level formed by the photolysis of ice mixtures containing deuterated methanol, CH2DOH or CH3OD, at 10 or 77 K. The analyses were performed using a state-of-the-art high-resolution mass spectrometer coupled with a compound-specific separation technique. The formation of d n -HMT (n = 0–8) was confirmed under all experimental conditions. In addition, methyl- and hydroxyl-substituted HMT and their deuterated isotopologues were also obtained in the products. The deuterium enrichment was outstanding when CH2DOH was used rather than CH3OD, and when photolysis was performed at 77 K rather than 10 K. We found that the deuteration level of the formed HMT far exceeded that of the reactants under the present experimental conditions. These results obtained during stable isotope probing of deuterium strongly suggest that HMT can play a role as an organic pool of interstellar D atoms. These may be distributed into other chemical species through molecular evolution in space.