The reactions are deduced to take place with the retention of the dimeric structures of the calcium hydrido‐alkyl and alkyl intermediates via a well‐defined sequence of Ca−H/C=C insertion and Ca−C hydrogenation events. This latter deduction is strongly supported by DFT calculations (B3PW91) performed on the 1‐hexene/H2 system, which also indicates that the hydrogenation transition states display features which discriminate them from a classical σ‐bond metathesis mechanism. In particular, NBO analysis identifies a strong second order interaction between the filled α‐methylene sp3 orbital of the n‐hexyl chain and the σ* orbital of the H2 molecule, signifying that the H−H bond is broken by what is effectively the nucleophilic displacement of hydride by the organic substituent.β‐Diketiminato (BDI) calcium alkyl derivatives undergo hydrogenolysis with H While related hydrogenation catalysis has also been extended to the reduction of more polarized imines,2H2(Me4TACD)2][BAr4]2  (2) in THF. The catalysis was proposed to proceed via linear calcium σ‐n‐alkyls that either rapidly cleave dihydrogen or themselves undergo β‐hydride elimination to reform the active calcium hydride species . Although 7–9 were stable in aliphatic solvents, solutions in benzene at even mildly elevated temperatures (ca. 50 °C) were found effect the unprecedented nucleophilic alkylation of a C−D bond of the C6D6 solvent. In this contribution, we further examine the reactivity of these heteroleptic calcium alkyls with dihydrogen and report our observations with regard to the activity of compound 3 for the catalytic hydrogenation of alkenes.In contrast, we have recently reported that a non‐THF solvated analogue of compound 6D6 solution of the calcium n‐hexyl derivative (9). Immediate analysis of the resultant 1H NMR spectrum confirmed that the solution comprised primarily a mixture of unreacted 9 (δ=4.69 ppm) and hydrogen (δ=4.46 ppm). Small quantities of the hydride (3) and the dicalcium n‐hexyl‐hydride (6), however, were also detected by their respective BDI methine resonances at δ=4.83 and 4.77 ppm. After storage at room temperature for 24 hours, the reaction was observed to contain a mixture of 3 and 9, whilst no significant increase in the intensity of the signals associated with the dicalcium hexyl‐hydride intermediate (6) was observed. The apparent steady‐state concentration of the dicalcium hexyl‐hydride suggested hydrogenation was occurring via a two‐step process, in which 6 is formed but more rapidly converted to the hydride (3). After 48 hours at room temperature, the complete conversion of 9 to 3 and the quantitative formation of n‐hexane were confirmed by the emergence of the BDI methine and hydride resonances of 3 (δ=4.83 and 4.27 ppm) and the triplet of n‐hexane (δ=0.89 ppm), which displayed a 1:1:6 ratio of intensities in the resultant 1H NMR spectrum. Although slower due to a likely primary kinetic isotope effect, analogous addition of D2 to a C6D6 solution of 9 similarly afforded 3‐2d and CH2D(CH2)3CH3, which could be clearly detected by the emergence of its BDI methine signal at δ=4.83 ppm in the resultant 1H NMR spectrum and by the relevant deuterated Ca−D and CH2D(CH2)3CH3 signals (δ=4.30 and 0.85 ppm) in the corresponding 2H NMR spectrum.An atmosphere of dihydrogen (2 bar) was added to a C3 with 1‐hexene under a dihydrogen atmosphere (2 bar) in C6D6 provided slow turnover at room temperature to hexane over the duration of three weeks as confirmed by the emergence of the diagnostic triplet signal of hexane at δ=0.89 in the resultant 1H NMR spectrum. Encouraged by this successful reactivity, the catalytic hydrogenation of a range of alkenes by 3 was investigated. The results of this study are summarized in Table An initial catalytic reaction mediated by 10 mol % of −1) and prolonged reactions times. This latter observation is interpreted to indicate that the slow rate of catalysis is a result of the limited solubility and availability of hydrogen in C6D6 under the conditions of the experiments (2 bar) rather than any variation in the nature of the substrates. At elevated temperatures, hydrogenation catalysis by 3 was suppressed by facile redistribution of the ligand system to the catalytically inactive homoleptic β‐diketiminate species, [(BDI)2Ca].1H NMR spectra throughout the hydrogenation of vinyltrimethylsilane, triphenyl(vinyl)silane, and norbornene . Notably, a significant quantity of the cyclized alkane, methylcyclopentane (56 %) was observed to form during the hydrogenation of 1,5‐hexadiene (Entry 9), which suggested the intramolecular cyclization is competitive with hydrogenation. In contrast, markedly less methylcyclopentane (4 %) was formed during Okuda's hydrogenation of 1,5‐hexadiene catalyzed by 2.6D6 solution of compound 3. Monitoring of the reaction at room temperature for 24 hours revealed the production of a mixture of unidentified products in addition to a dicalcium cyclopentylmethyl‐hydride species (10) and the corresponding dicalcium‐dicyclopentylmethyl derivative (11), identified by the emergence of BDI methine and upfield methylene doublet signals with relative intensities of 1:2 at δ=4.72 and −0.55 ppm, respectively. Despite the mixture of products formed, single crystals deposited from the reaction mixture and the resultant X‐ray diffraction analysis allowed the structure of 11 to be confirmed as the result of facile carbocalciation and 5‐exo‐trig cyclization  at room temperature, albeit with similar turnover frequencies (0.02–0.04 h1 was reported to occur over a shorter timeframe (17–25 hours) than the corresponding results shown in Table 2H2(Me4TACD)2][B(C6H4‐4‐tBu)4]2 (2) significantly expanded this substrate scope to include less activated terminal alkenes.2, however, also ensued without any observable formation of the necessary alkyl intermediates, an observation which was ascribed to their instantaneous consumption either by hydrogenolysis or competitive β‐hydride elimination  under 1 atmosphere of 1‐butene by 1H NMR spectroscopy demonstrated that the 1‐butene was only very slowly consumed over the course of four weeks. This process occurred with the simultaneous emergence of a new alkene multiplet at δ=5.48–5.53 ppm and an aliphatic doublet signal at 1.55 ppm, which were assigned to the generation of the internal alkene, 2‐butene. Although not investigated further, the formation of 2‐butene is proposed to have arisen from an effective chain walking mechanism involving the β‐hydride elimination reaction of 9, 2,1‐insertion of a molecule of 1‐butene into the reformed Ca−H bond and a subsequent β‐hydride elimination reaction.The hydrogenation of the activated alkenes, 1,1‐diphenylethylene, styrene and α‐methylstyrene catalyzed by compound 3. The mechanism is again predicated upon a cascade of polarized insertion reactions in conjunction with irreversible hydrogenolysis and occurs with the maintenance of dimeric calcium species throughout the catalysis. The stoichiometric reactivity studies described above indicate that insertion of alkenes occurs in a stepwise fashion, whereupon the resultant dicalcium alkyl‐hydride intermediates are produced in an effective steady state throughout the course of catalysis. Although these latter species have the potential to undergo a second alkene insertion, we suggest that turnover is largely based on their interception and hydrogenolysis by H2.On the basis of these observations, the mechanism shown in Scheme n‐hexyl derivative (9), through the reaction of compound 3 with 1‐hexene.3 was described in our previous report and the associated barrier yielding the dicalcium n‐hexyl‐hydride complex C (6) is 20.1 kcal mol−1. From C (6), either hydrogenation by H2 .Further insight into this proposed mechanism was provided by density functional theory (DFT) calculations carried out with the same computational approach  previously used to describe the stepwise formation of the dicalcium di‐−1) of C (6) and 1‐hexene insertion  is 4.0 kcal mol−1, indicating a marginal preference for the direct hydrogenation. Within the precision of the method, however, the possibility of 1‐hexene insertion cannot be ruled out completely. The complete hydrogenation subsequent to the second 1‐hexene insertion, that is, from the dimeric n‐hexyl complex F (9), is also reported on Figure S21. The two barriers found for the hydrogenation from F (9)  are of a similar magnitude as the barrier for the hydrogenation from C (6). The hydrogenation, therefore, may be deduced to be similarly kinetically accessible for both possible pathways.The difference between the barrier heights associated with the direct hydrogenation  is clearly indicative of a heterolytic cleavage of the H2 molecule. The HOMO−2  involves the filled sp3 orbital of the n‐hexyl chain, the H2 σ* and a 3d orbital of both Ca centers. The involvement of 3d orbitals on Ca has recently been invoked as a potentially significant factor in the structural and reaction chemistry of organocalcium species.n‐hexyl unit is better viewed as a proton, whereas the hydrogen atom interacting with the two Ca centers is much more hydridic in nature. This interpretation is further corroborated by the corresponding NBO analysis in which a strong second order interaction is observed between the filled α‐methylene sp3 orbital of the n‐hexyl chain and the σ* orbital of the H2 molecule, signifying that the H−H bond is broken by what is effectively the nucleophilic displacement of hydride by the organic substituent. This electron density is then redistributed via by a less strong second order interaction between the σ*H−H orbital and the two Ca atoms. The highly polarized nature of this process is underscored by the NPA charge analysis, which indicates a charge of −1.0 on the α‐methylene carbon of the n‐hexyl chain, charges of +1.50 and +1.43 for the two Ca centers and charges of −0.23 and +0.03 for the two H2 hydrogen atoms. An effectively identical interpretation also emerges for the hydrogenation TS from the dicalcium di‐n‐hexyl species F (9) .The hydrogenation transition states are particularly notable and display features which discriminate them from a classical σ‐bond metathesis. Examination of the relevant molecular orbitals and the NBO analysis of the hydrogenation of the In conclusion, hydrogenation of the Ca−C bonds of β‐diketiminato calcium alkyls occurs through retention of the dimeric calcium alkyl and hydrido‐alkyl species allowing the catalytic hydrogenation of a wide range of olefinic substrates under mild conditions. The hydrogenolysis step ensues through an unprecedented bimetallic nucleophilic pathway in which the H−H bond is cleaved heterolytically, that was so far only reported on monometallic species.The authors declare no conflict of interest.As a service to our authors and readers, this journal provides supporting information supplied by the authors. Such materials are peer reviewed and may be re‐organized for online delivery, but are not copy‐edited or typeset. Technical support issues arising from supporting information (other than missing files) should be addressed to the authors.SupplementaryClick here for additional data file.


With an ultralarge surface-to-volume ratio, a recently synthesized three-dimensional graphene structure, namely, carbon honeycomb, promises important engineering applications. Herein, we have investigated, via molecular dynamics simulations, its mechanical properties, which are inevitable for its integrity and desirable for any feasible implementations. The uniaxial tension and nanoindentation behaviors are numerically examined. Stress–strain curves manifest a transformation of covalent bonds of hinge atoms when they are stretched in the channel direction. The load–displacement curve in nanoindentation simulation implies the hardness and Young’s modulus to be 50.9 GPa and 461±9 GPa, respectively. Our results might be useful for material and device design for carbon honeycomb-based systems.  Graphene is known widely, due to its excellent mechanical nature, as a so-called “miracle material”, with many of its characteristics measured experimentally or theoretically exceeding those obtained in other materials: A Young’s modulus of 1 TPa and intrinsic strength of 130 GPa , high stn CHC) and zigzag CHC (zzm CHC) [n or m in zzm) varies, the cell sizes of CHCs and, thus, their mechanical properties change correspondently. Krainyukova et al. [m–sp2–sp3 CHC structures and revealed that the carbon modifications are as stable as diamond by solid-state semi-empirical SCF methods using the modified neglect of diatomic overlap (MNDO) Hamiltonian [m–sp2–sp3 CHCs with different m  using ab initio pseudopotential as well as the environment-dependent tight-binding method, making clear that the carbon allotrope is elastically stable and has a fairly high shear modulus [n–sp2–sp3 CHCs with different cell sizes via molecular dynamics simulation using optimized reactive empirical bond-order potential [2–sp2–sp3 CHCs, ac3–sp2–sp3 CHCs, and zz2–sp2–sp3 and zz2.5–sp2–sp3 CHCs [n–sp2 and zzm–sp2–sp3 CHCs via MD analysis with the Adaptive Intermolecular Reactive Empirical Bond Order potential (AIREBO) [n–sp2–sp3 and zzm–sp2–sp3 CHCs using MD simulation with AIREBO potential [2–sp2–sp3 CHC to an ac2–sp2 CHC, which causes the change of tensile deformation behavior of ac2 CHCs. In the meanwhile, we gave the stress–strain curves, Young’s modulus, and Poisson’s ratio of ac2 CHCs, which show a great agreement with previous studies. We also built an out-of-plane nanoindentation test to investigate the plastic deformation behaviors of ac2–sp2–sp3 CHCs, giving the hardness and Young’s modulus. According to the cell patterns in the plane perpendicular to the cell axis, carbon honeycombs can be categorized into two sets: Armchair CHC (aczzm CHC) . When tha et al.  synthesia et al. ,22,23,24otential . Gu et asp3 CHCs . Zhang esp3 CHCs  built th(AIREBO)  potentiaotential . Despite3 or sp2 bonding  or one variant of diamond-like sp3 bonds with both adjacent hinge and non-hinge atoms. In the case of sp2 bonds, hinge atoms do not interact with each other, allowing for electron exchange with only adjacent tube atoms. In the case of sp3 bonds, it is not only nearby tube atoms but also nearest-neighbor hinge atoms that get bonded with the center hinge atom, resulting in a diamond-like atom-hinge that ties up three around the atomic walls. According to the carbon patterns in the plane perpendicular to the axis of cell channel, carbon honeycombs can be categorized into two sets: Armchair CHC (acn CHC) and zigzag CHC (zzm CHC) [2 and sp2–sp3 CHCs. In zzm CHCs, carbon atoms of graphene-like walls are bonded by sp2 bonds, while hinge atoms can be bonded with adjacent atoms only by sp3 bonds. Therefore, zzm CHCs always refer to zzm–sp2–sp3 CHCs. However, the other case is different. In acn CHCs, hinge atoms can be bonded with atoms around them by sp2 or sp3 bonds. Therefore, acn CHCs can be subdivided into acn–sp2 CHCs and acn–sp2–sp3 CHCs. In the case of acn–sp2–sp3 CHCs, hinge atoms can be distributed symmetrically or asymmetrically in a crystal cell, according to which acn–sp2–sp3 CHCs can also be subdivided into a symmetrical one and asymmetrical one, as shown as Carbon honeycomb, just as its name implies, is a kind of tubular structure with a honeycomb-like pattern through a top–down perspective. The tube wall can be regarded as a graphene-like monolayer, thereby deemed to be a 3D version of graphene. One thing about its structure that remains to be discussed is whether three adjacent tube walls are bound up with each other via sp bonding b,c. Undozzm CHC) , as show3–sp2 and sym-ac3–sp2–sp3 CHCs, as shown in 3–sp2–sp3 CHC, stronger sp3 bonds between the hinge atom and nearest-neighbor tube atoms can be observed than those between two adjacent hinge atoms, the bond angle of which is measured as 103.35°. Meanwhile, exactly strong sp2 bonds among graphene-like atoms can be distinguished from sp3 bonds, with denser electron density around tube atoms observed  calculation for both primitive cells of sym-ac3–sp2 CHC and sym-ac3–sp2–sp3 CHC. The reason we focused on them is that in the work of Krainyukova et al. [3–sp2 CHC according to their synthetic CHC samples, proving that ac3 CHCs are more likely to exist in the nature. The method of MD simulations is well established to investigate the elastic and plastic behaviors of sym-ac3–sp2 and sym-ac3–sp2–sp3 CHCs. Initially, we carried out uniaxial tension tests for both types of CHCs, simultaneously determining the Young’s moduli of the sym-ac3–sp2–sp3 CHC to be 551 ± 4 GPa and of sym-ac3–sp2 CHC to be 542±4 GPa in the channel direction  and zero pressure in the NPT ensemble  using AIREBO [−1. Considering that temperature effect will cause random distribution of atoms, which will prevent us from observing the bonding transformation of hinge atoms, in addition to at room temperature, uniaxial tension tests were also carried out at 10K, at which little thermal motion is allowed so that a perfect configuration might remain. We used a molecular dynamics (MD) software, LAMMPS , which ig AIREBO  potentia3–sp2–sp3 CHC along the channel direction. A thick CHC plate measured as 17.82 nm × 15.43 nm × 11.60 nm was set up in a simulation box with a vacuum layer 9.00 nm thick in addition . From a previous study [From uniaxial tension, we obtained the engineering strain–stress curves of both the sym-ac–sp3 CHC , which cus study , the You3–sp2–sp3 CHC deviates a little from that of the sym-ac3–sp2 CHC. Then, in a certain strain range (from 0.072 to 0.132 for case at 10 K), a yield stage occurs in the deformation of the sym-ac3–sp2–sp3 CHC, which indicates a plastic deformation has initiated. Later in this article, the mechanism therein will be discussed in detail. After the yield step, the sym-ac3–sp2–sp3 curve behaves just like a sym-ac3–sp2 curve does, even breaking off at the same fracture strength, which is around 61 GPa (56 GPa for case at 300 K), with a retardation measured as around 0.06 in strain for the case at 10 K, equal to the length of the preceding yield stage. Thus, the mechanical behavior of the sym-ac3–sp2–sp3 CHC can be distinguished from that of the sym-ac3–sp2 CHC by the fracture retardation caused by yield terrace mentioned above. When the tension starts, both CHCs are stretched quasilinearly under homogeneous and elastic deformation; herein, the initial curve of the sym-ac3 bond, the neighboring hinge atoms must be bonded with each other, thus exhibiting a high channel-direction stress; while for the case of sp2, hinge atoms do not interact with each other, so there will be a low and even zero channel-direction stress on them. In addition, sp2 bonding is in-plane , while sp3 is three-dimentional; thus, we can also tell them apart by their geometry and nearest neighbors. Three snapshots at different states of strain, corresponding to three characteristic points in 3 bond between two hinge atoms, while the sp3 configuration remains. However, when the engineering strain reaches around 0.072, some sp3 pairs of hinge atoms break up, the distortion energy of graphene-like atoms begins to increase gradually, and simultaneously, the yield stage starts. When the strain attains 0.132, almost all sp3 pairs are disconnected, most of the deformation is shared by the atomic cellular walls, and the yield stage terminates at the same time. By contrast, the sym-ac3–sp2 CHC, without sp3 interaction, does not experience such a transformation, and atomic tubes instead of hinge atoms bear most proportions of tensile force. Compared with graphene, although CHC has a lower fracture strength and modulus, it can obtain a higher fracture strain than graphene through this transformation, which may be applied in flexible coating. In order to illustrate the mechanism of this yield stage, a slice through some sheets of graphene-like walls has been exhibited in At present, nanoindentation is commonly used for the research of mechanical properties of materials on the nanoscale ,31,32. Tn–sp2 and zzm–sp2–sp3 CHCs were reported. However, the indent depth was small (4 Å) and the unloading process was not displayed. Herein, we simulated the process of nanoindentation in the channel direction of the sym-ac3–sp2–sp3 CHC, an indentation depth of about 40 Å, containing two steps, acting load and unloading, via LAMMPS [In a previous study , nanoinda LAMMPS , under tP and projected contact area pA [Nanoindentation hardness is the average pressure the material can support under an external load, defined as the ratio of indentation load  area Ap . From thrE, which can be expressed as: S is termed as contact stiffness, which can be obtained as the slope at the tip of unloading curve, β is a constant that depends on the geometry of indenter, for a spherical indenter, β = 1. Furthermore, we can determine the substrate’s modulus using following relationship that reflects impacts of both indenter’s and substrate’s moduli:i refers to indenter, while s refers to substrate. In addition, Young’s modulus can also be calculated indirectly via reduced modulus 3–sp2–sp3 CHC in the process of nanoindentation, five snapshots are displayed in The nanoindentation hardness is determined as 50.9 GPa in 3–sp2 CHCs and especially the sym-ac3–sp2–sp3 CHC which is slightly more stable according to our DFT calculation and previous experimental study, using LAMMPS to carry out a typical uniaxial tension and nanoindentation simulation. In the case of uniaxial tension, we discussed the yield stage in the stress–strain curve of the sym-ac3–sp2–sp3 CHC caused by a intriguing transformation of covalent bonds of hinge atoms from strong sp3 to comparatively weak sp2, and we also determined the Young’s moduli at room temperature, 542 ± 4 GPa for the sym-ac3–sp2 CHC and 551 ± 4 GPa for the sym-ac3–sp2–sp3 CHC, which are in great agreement with a previous analytical study. Then, in the discussion of nanoindentation, we determined the hardness (50.9 GPa) and Young’s modulus (461 ± 9 GPa) via the load–displacement curve and gave an insight into the plastic deformation caused by the indenter on the nanoscale, which can be explained by the permanent conversion of covalent bonds from sp3 type to sp2 type. These findings provide an insight into the relationship between the covalent bond type of hinge atoms in carbon honeycomb and the plastic behavior of carbon honeycomb. We discussed the elastic and plastic behaviors of sym-ac


The influence of rapid thermal annealing (RTA) treatment on the sensing and impedance properties of the YbTaO4 sensing membranes deposited through reactive co-sputtering onto Si substrates was explored. X-ray diffraction, atomic force microscopy, and X-ray photoelectron spectroscopy revealed the structural, morphological, and chemical features, respectively, of these YbTaO4 films annealed at 700, 800 and 900 °C. The YbTaO4 EIS device annealed at the 800 °C exhibited a super-Nernstian response of 71.17 mV/pH within the pH range of 2–12. It also showed the lowest hysteresis voltage ( < 1 mV) and the lowest drift rate (0.22 mV/h) among the tested systems. Presumably, the optimal annealing temperature improved the stoichiometry of YbTaO4 film and increased its (−131)-oriented nanograin size. Moreover, the impedance properties of YbTaO4 EIS sensors were investigated by using the capacitance–voltage method. The resistance and capacitance of YbTaO4 sensing films annealed at three various temperatures were evaluated by using different frequency ranges in accumulation, depletion, and inversion regions. The semicircle diameter of the YbTaO4 EIS sensor became smaller, due to a gradual decrease in the bulk resistance of the EIS device, as the RTA temperature was increased.In this study we developed ytterbium tantalum oxide (YbTaO Over the past four decades, research on various kinds of ISFET-based biosensors, for example glucose, urea, protein, DNA hybridization, and DNA methylation detection7, has been conducted because they have the advantages of fast response, small size, and low cost. In general, the gate electrode of a conventional metal-oxide-semiconductor field-effect transistor (MOSFET) is replaced by a chemically sensitive oxide layer, an electrolyte, and a reference electrode to become an ISFET device. An electrolyte-insulator-semiconductor (EIS) capacitor has a simple structure and easy fabrication with respect to an ISFET device. The surface potential of such an ISFET is modulated by the change in the charges at the interface between the oxide layer and electrolyte, thus leading to the shift of the threshold voltage and the variation of the drain-source current. As a result, quality and chemical stability of the sensing film play a key role in the applications of an ISFET or EIS device. The most widely used sensitive oxide film is SiO2 in ISFETs, but its application is severely limited by its poor sensitivity8. Therefore, Si3N4, Al2O3, Ta2O5, TiO2, ZrO2, and HfO2 films were investigated as alternative sensitive oxides in ISFETs or EISs to improve their sensitivities12. However, the pH sensitivity of the traditional ISFET or EIS sensors could not exceed the Nernstian limit (59.18 mV/pH at 25 °C) because of intrinsic properties of these metal oxide films. In addition, there are some materials related problems with silicide at the interface of oxide film/Si substrate after thermal treatment and dangling bonds on the sensing film during the thin-film deposition to impact their sensing performance14.An ion-sensitive field-effect-transistor (ISFET) device was first developed by Bergveld in 1970 as a replacement for a fragile glass electrode16. Of these RE oxides, ytterbium oxide (Yb2O3) thin film turns out to be a potential gate oxide because of its novel properties, including excellent thermal and chemical stability, large bandgap (~5 eV), and high κ (~15), in which the κ values depend on deposition processing18. Nevertheless, due to the highly hygroscopic nature, there is a need to develop the fabrication of stable RE oxide film, suppressing the formation of a hydroxide layer on the film because of the presence of oxygen vacancies19. The formation of oxygen vacancies can contribute to the occurrence of a sufficient atomic reorganization in the film structure. In order to solve the aforementioned shortcoming, in an effort to remove the oxygen vacancies in the RE oxide film, the addition of Ti or TiO2 into the film could be reduced the moisture absorption of RE oxides, thereby improving the structural and electrical properties21. Recently, our group previously demonstrated the structural and sensing characteristics of Yb2Ti2O7 sensing films in an EIS sensor22, in contrast, the resistance to the level of moisture absorption for the incorporation of Ta into the Yb2O3 film after post-annealing treatment to eliminate the oxygen vacancies is still unclear. The aim of this paper is to explore the effect of post-annealing treatment on the structural, sensing, and impedance characteristics of ytterbium tantalum oxide (YbTaO4) sensing films deposited on Si substrates through reactive rf co-sputtering. X-ray diffraction (XRD), atomic force microscope (AFM), and X-ray photoelectron spectroscopy (XPS) were employed to examine the film structures, surface morphologies, and chemical compositions of YbTaO4 films annealed at three different temperatures , respectively. In addition, structural characteristics of the YbTaO4 films are correlated to sensing and impedance properties of the EIS sensors after annealing at three temperatures. In this study, the YbTiO4 membrane after RTA at 800 °C showed a higher pH sensitivity (71.17 mV/pH), a smaller hysteresis voltage (<1 mV) and a lower drift rate (0.22 mV/h), compared with other RTA temperatures.In search of the low defect density and high thermal stability in high dielectric constant (κ) metal oxide films, rare-earth (RE) oxide thin films have been studied for use as a replacement MOSFET gate dielectric due to their high κ values, large bandgap energies, good thermodynamic properties, high resistivities, and high conduction-band offsetsxOy thin films were grown on p-Si (100) substrates with a resistivity of 5–10 Ω-cm by rf co-sputtering using from metal Yb and Ta as target materials in a mixture of Ar/O2 (5 sccm/20 sccm). The plasma power of Yb and Ta targets was 100 W. The chamber pressure was 1 × 10−3 Torr during the growth process and the substrate temperature was 27 °C. The growth rate and physical thickness of the YbTaxOy film were ~2 nm/min and ~60 nm, respectively. Subsequently, the samples were annealed at three various temperatures  by rapid thermal annealing (RTA) in oxygen (O2) ambient for 30 s to form an YbTaO4 compound. Next, the back-side oxide of Si wafer was etched by buffer oxide etchant (BOE) solution and Al (400 nm thick) as a back-side electrode was deposited by thermal evaporation to achieve good electrical contact. The sensing area of 3.14 mm2 was defined by a robotic dispensing system using an adhesive silicone gel (S181) as an isolating layer. The EIS device was mounted on the Cu-coated printed circuit board (PCB) with an Al back-side contact through Ag paste. Finally, the non-sensing region of the EIS sensor was covered with epoxy.Prior to the deposition of sensing film, the Si substrate was cleaned and hydrogen-terminated by using diluted HF. The YbTaα radiation (λ = 1.5406 Å) was run under a voltage of 40 kV and a current of 20 mA. A tapping mode AFM (NT-MDT Solver P47) was employed to explore the surface topography and determine the surface roughness  of the YbTaO4 films annealed at three temperatures. The Rrms roughness of these samples was estimated in scanning areas of 3 × 3 μm2. The film composition of YbTaO4 films was analyzed using XPS (Thermo VG Scientific Microlab 350 system) with a monochromatic Al Kα source (1486.7 eV). The binding energy scale of each element was calibrated by setting the main hydrocarbon peak at a binding energy of 285 eV (C 1 s).The orientation and phase of these films were investigated by XRD (Bruker D8 discover diffractometer) in a scan range of 2θ = 20°–60° using a step time of 1 s and a step size of 0.05°. The Cu K4 EIS devices were evaluated by capacitance–voltage (C–V) curves. The C–V measurements for different pH buffer solutions (Merck Inc.) were performed using inductance–capacitance–resistance (LCR) meter (Agilent 4284 A), operated at 500 Hz with an applied ac voltage of 10 mV. The Ag/AgCl electrode  electrode as a reference electrode was used. The impedance measurement of EIS devices was performed by a combination of Agilent 4284 A and 4285 A LCR meters. To achieve the steady results, all samples were kept in reverse osmosis (RO) water for 24 h before measurement. The sensing membrane was washed with deionized water before transferring to subsequent pH solution. In order to prevent light and noise interference, all the measurements were performed in a dark box. Three devices on each sample were repeated at least three times to measure sensing performance of the YbTaO4 EIS sensors.The pH sensitivity, hysteresis voltage, and drift rate of the YbTaO4 films deposited on a Si substrate after RTA at different temperatures (700–900 °C) in O2 ambient were presented in Fig. 4 films after RTA at three temperatures have a monoclinic structure. The YbTaO4 diffraction peaks were recorded at (310) and (−131) peaks for the film annealed at 700 °C. Apart from the increment in the intensity of the existing YbTaO4 peaks, an additional YbTaO4 (011), (−111), (111), (020), (200), (120), (022), and (202) peaks (JCPDS: 00-024-1416) were detected when the annealing temperature was raised up to 800 °C, suggesting a polycrystalline structure. These peaks became more noticeable as the annealing temperature increased. By further increasing the annealing temperature to 900 °C, the increment in the intensity of YbTaO4 (−111), (111), (020), (200), and (120) peaks was observed. Overall, as the RTA temperature increased from 700 °C to 800 °C, the intensity of the YbTaO4 (−131) peak increased. This condition may indicate the reduction of oxygen vacancies present in the film due to the highest stability offered by (−131)-oriented YbTaO4. The crystallite size (or grain size) of the YbTaO4 film after three RTA temperatures is determined by using the Scherrer’s equation for the main diffraction peak23. The average grain size was calculated to be 0.12, 0.21, and 0.24 nm for the sample annealed at 700, 800 and 900 °C, respectively. Moreover, the crystallite size of the (−131) peak for the YbTaO4 film annealed at 700, 800 and 900 °C was 0.12, 0.35 and 0. 19 nm, respectively.The XRD patterns of all YbTaO4 thin films deposited on a Si substrate was evaluated by AFM to examine the effect of RTA temperatures. Figure rms roughnesses of the YbTaO4 films annealed at various temperatures. In general, the sample is uniform and smooth without defects, such as a crack or void. It can be observed in Fig. rms roughness of the YbTaO4 films annealed at 700, 800 and 900 °C was 0.51, 0.69 and 0.83 nm, respectively. The Rrms roughness of the sample is in the increasing trend as the RTA temperature increases. The reason of increase of surface roughness might be attributed to the growth of grain size due to the formation of grain agglomerations in the film after annealing at high temperature in oxygen gas. This result indicates that the grain size increases with increasing RTA temperature, as expected, leading to the enhanced crystallinity and the sharpened diffraction peaks. Therefore, the obtained Rrms roughness from AFM matches with the XRD results. Hence, more YbTaO4 was formed, and a noticeable change in the surface morphology was observed for the sample annealed at 900 °C.The surface morphology of YbTaO4 layer were analyzed by XPS with Ar+ ion etching. The 30 s etching process was conducted to remove the surface contamination. The obtained Yb 3d, Ta 4 f and O 1 s spectra of the YbTaO4 films after RTA at three temperatures were shown in Fig. 4. The ytterbium compounds have two valence states. The filled 4 f shell of divalent ytterbium (Yb2+) in the 4d spectrum is a doublet with a peak area ratio of 3:2, while partially filled 4 f shell of trivalent ytterbium (Yb3+) is a multiplet24. The electronic structure of the Yb 4d multiplet peaks in these films is in good agreement with multiplet splitting in the Yb 4d spectrum of Yb2O3, which is different from the 4d doublet of Yb2+. There are two main Yb 4d3/2 and 4d5/2 peaks centered at 199.5 ± 0.1 eV and 185.3 ± 0.1 eV, respectively, which are consistent with the binding energy of Yb 4d for Yb2O3 ref.25. The Yb 4d5/2 peak of the film annealed at 800 and 900 °C was slightly shifted toward a higher binding energy, suggesting the incorporation of more Ta atom into the Yb2O3 film forming a YbTaO4 compound. Figure 4 films annealed at three temperatures. The Ta 4f5/2 and 4f7/2 peaks located at binding energies of 28.1 and 26.2 eV, respectively, are assigned to pure Ta2O5 film26. The spin-orbit splitting energy of Ta2O5 film was 1.9 eV from Ta 4f5/2 to Ta 4f7/2 peak. The deconvolution results of these YbTaO4 films demonstrated a perfect fit for Ta 4f5/2 and 4f7/2 peaks located at two 28 ± 0.1 and 26.1 ± 0.1 eV and splitting energy of ~1.9 eV, which are in good agreement with the reported values for Ta2O5 ref.26 indicating that the Ta exhibits the highest oxidation state. These sensing films consist of tantalum suboxides (Ta2O5) with no TaOx content. The splitting energy of about 1.9 eV can consider strong binding interaction between the tantalum and the oxygen atoms. The reaction of Ta2O5 with Yb2O3 can form a YbTaxOy compound. The O 1 s XPS spectra of the YbTaO4 films after RTA at different temperatures were deconvoluted with Gaussian-Lorentzian curve fitting after the Shirley background, as shown in Fig. 2− species occurring in Ta2O526. The low binding-energy peak of 529.8 eV can be attributed to the Yb-O-Ta bonding. These low peak intensities are assigned to O-C (or O=C) bonds resulting from various species present in the organic carbon overlayer on top of the YbTaO4 films. The peak intensities of YbTaO4 and Ta2O5 components for the film annealed at 800 °C exhibited higher and lower, respectively, in the O 1 s signal compared with other temperatures. In addition, the O 1 s peak corresponding to YbTaO4 for the sample annealed at 900 °C was a lower intensity than that at 800 °C. During high-temperature annealing, the more Ta or/and Yb atoms diffuse readily from the YbTaO4 film to form a thicker silicate layer at the YbTaO4–Si interface15.The valence states of Yb, Ta, and O elements in a YbTaO4 EIS sensors after RTA at three temperatures was investigated. The flatband voltage (VFB) of an ISFET or EIS device can be expressed as:27Eref is the reference electrode potential, ψ0 is the surface potential, χsol is the surface dipole potential of the solution, ΦSemi is the semiconductor work function, q is the elementary charge, Qeff is the effective charge produced at the oxide-semiconductor interface and in the oxide by the different types of charges , and Cox is the gate oxide capacitance. Apart from ψ0, all these terms are fixed values. This term causes an ISFET or EIS device responsive to the tested solution as a result of the polarization and formation of the potential barrier, which is related to the H+ concentration. Therefore, an ISFET or EIS device can be sensitive to the solution pH. The potential of electrolyte-insulator interface corresponding to pH can be explained by using a combination of the site-binding model and the Gouy–Chapman–Stern theory28. The pH sensitivity of an ISFET device is derived from Bergveld28 as follows:α represents a dimensionless sensitivity parameter which varies between 0 and 1, Cdiff indicates the differential capacitance of the electric double-layer, and βint stands for the intrinsic buffer capacity of the oxide surface. The above equation gives information that the βint and Cdiff are effectively influenced the pH sensitivity of an ISFET device. The Cdiff is obtained from the Gouy-Chapman-Stern model. In addition, the βint is related to the density of binding sites on the gate oxide. The higher the intrinsic buffer capacity βint is, the higher the pH sensitivity of the sensing film will achieve. The intrinsic buffer capacity is associated with the surface roughness and the material quality of the film. The change in the pH value of the electrolyte solution could give rise to a shift of the flatband voltage in the C-V curves.After the film material analyses, the sensing performance  of YbTaO4 EIS sensors after RTA at different temperatures, a set of C-V curves measured in a wide range of pH 2–12 was tested. Figure 4 EIS devices annealed at 700, 800 and 900 °C, respectively. For a p-type Si substrate, three zones were visible, namely the accumulation, inversion and depletion regions. The accumulation region is caused by a hole channel on the Si surface when a high positive voltage is applied to substrate electrode. In contrast, when a high negative voltage is applied to substrate electrode, an inversion layer of electrons is formed in the inversion region. The C-V measurements around the depletion region were conducted for electrolyte solutions with pH ranging from 2 to 12. It is evidence that the kinks of the EIS sensors annealed at 800 and 900 °C were found in the depletion region of the C-V curves, possibly indicating the presence of interface state at the oxide-substrate29. In an ISFET operation, an adequate number of surface hydroxyl (OH) groups is essential on the gate oxide material for pH measurement at the Nernstian limit. The OH groups can protonate  or deprotonate 27, which relies on the solution pH value. Therefore, the surface potential of the gate oxide changes, which can be measured via a capacitance variation of an EIS device. A negative shift of the reference voltage (VREF) with increasing pH values of the investigated electrolyte solution reflects a more negatively charged gate oxide surface. The inset of Fig. 4 EIS devices after RTA at 700, 800 and 900 °C, respectively. The 0.5Cmax was set as the reference to extract the VREF versus the change of the pH value. The pH sensitivity and linearity of the C-V curves were achieved by linear regression. The YbTaO4 EIS device annealed at 800 °C exhibited the highest sensitivity of 71.17 ± 2.36 mV/pH among these RTA temperatures (52.37 ± 3.25 mV/pH for 700 °C and 61.13 ± 2.26 mV/pH for 900 °C). This result is mainly attributed to the film with a stoichiometric YbTaO4 structure and a larger grain size of (−131) plane enhancing the pH sensitivity. Furthermore, the optimal RTA temperature in the oxygen ambient might improve the surface and interfacial material quality and increase the intrinsic buffer capacity (βint). Furthermore, the pH sensitivity of the YbTaO4 EIS sensor is higher than the theoretical Nernstian value of 59.4 mV/pH at 27 °C. This super-Nernstian value of our EIS sensor could be related to the mechanism of one transferred electron per 1.5 H+ ion30. The decrease in pH sensitivity after RTA at 900 °C is the decrease of the surface state density of the OH groups31.In order to evaluate pH sensitivity of the YbTaO4 EIS sensors with three RTA temperatures were shown in Fig. 32. The YbTaO4 EIS device after RTA at 800 °C had the lowest hysteresis voltage of 1 ± 0.2 mV among these annealing temperatures (63 ± 8.9 mV for 700 °C and 11 ± 2.8 mV for 900 °C), suggesting that the optimal RTA temperature could effectively reduce the oxygen vacancies and dangling bonds. On the contrary, the EIS sensor annealed at 700 °C showed a relatively larger hysteresis voltage of 63 mV compared to other temperatures. This result could be attributed to a high number of defects in the YbTaO4 film because these defects cannot be removed by the low annealing temperature. Figure 4 sensing films annealed at three RTA temperatures. Each of the EIS sensors was measured in a pH 7 buffer solution for a period of time. The change in the VREF can be expressed as ΔVREF = VREF(t)–VREF(0). The slope of the drift characteristics indicates the drift rate of an EIS device. The drift effect can be interpreted by the hopping and/or trap-limited transport of water-related species33, which the localized defects could interact with the tested solution, thus resulting in the gate voltage shift. Figure 4 EIS sensor with the 800 °C had the lowest drift rate of 0.22 ± 0.03 mV/h, whereas the EIS device with the 700 °C featured the highest drift rate of 0.36 ± 0.08 mV/h. The lower drift rate may be due to the fact that the crystal defects could be eliminated by optimal RTA temperature in O2 ambient, hence causing a lower capacitance of hydrated layer. In contrast, the higher drift rate could be contributed to the higher capacitance value of hydrated layer. In Table 4 membrane is compared with commonly used materials for EIS or ISFET-based sensors such as Ta2O59, Al2O310, ZrO211, HfO211, TiO234, SnO235, and Yb2Ti2O722. It is found that our YbTiO4 membrane demonstrated a higher pH sensitivity (71.17 mV/pH), a smaller hysteresis voltage (<1 mV) and a lower drift rate (0.22 mV/h), relative to those of these materials.Furthermore, the hysteresis curves of the YbTaO4 EIS sensor annealed at 800 °C for various H+, K+, Na+, Ca2+, and Mg2+ ion concentrations in pH 7. The response curves were measured in these ion concentrations ranging from 10−5 to 0.05 M. It is clear that the YbTiO4 sensing membrane had high selectivity to H+ ions and less selectivity to other ions, e.g. K+, Na+, Ca2+, Mg2+.Figure 4 EIS devices annealed at three different RTA temperatures and tested at different solution pH values were presented in Figs 4 EIS devices demonstrated a depressed semicircle portion at high frequency region and a slanted straight portion at low frequency region. The results indicate that the semicircle portion is related to charge transfer-limited process and the linear portion is associated with diffusion limited process or mass transfer process at oxide–electrolyte interface. At higher frequency end the intercept corresponds to the Si substrate impedance (ZS) and at lower frequency corresponds to the sum of ZS and charge transfer resistance (Rct). The value of Rct is a measure of electron transfer across the exposed area of the oxide surface. The conductance method, indicating the loss mechanism because of interface trap capture and emission of carriers, is generally employed to evaluate the density of interface trap states (Dit) in the depletion region for a MOSFET device29. If the capacitance has small losses in the oxide, a simplified equivalent circuit of an EIS sensor for conductance method was shown in Fig. 4 EIS sensors with different RTA temperatures from 700 to 900 °C.Electrochemical impedance spectroscopy is well established as a powerful tool for investigating the mechanisms of electrochemical reactions and measuring the dielectric and transport properties of materials. In addition, it can be successfully applied for the characterization of biosensing surfaces and/or in estimation of bioanalytical signals produced by biosensors. A small sinusoidal perturbation of potential measured is usually applied to an electrochemical device at different frequencies, thus monitoring the variation of electric current or voltage. Figure 4 EIS sensors annealed at 700 °C exhibited larger compared with other temperatures, as shown in Fig. 4 EIS sensors after RTA at three temperatures were performed under three regions and then tested at pH 7. Here, a lower real impedance value  occurred at higher frequency, whereas a higher real impedance value (large semicircle) appeared at lower frequency. The impedance spectra (Zim vs. Zre) of the YbTaO4 EIS sensors annealed at the three temperatures and then tested at pH 10 under the three regions were shown in Fig. 3O+ ions is larger than that of the HO− ions, thus leading to a low diffusion rate. Therefore, the bulk resistance of the EIS devices gradually increased from ~0.4 to ~1 kΩ as the pH value increased.At pH 4, a semicircle diameter of the YbTaO4 sensing films on Si substrates by means of reactive rf cosputtering was explored in the paper. Material analyses indicate that the YbTaO4 sensing film after RTA at 800 °C could form a stoichiometric YbTaO4 structure and increase the grain size of (−131) plane. The YbTaO4 sensing film annealed at 800 °C exhibited a higher pH sensitivity of 71.17 mV/pH, a smaller hysteresis voltage of 1 mV and a lower drift rate of 0.22 mV/h, in comparison with other RTA temperatures. These results are attributed this RTA temperature to the reduction in the defects and the improvement in the material quality of the film and the interface of YbTaO4-Si substrate. The semicircle radius of the Nyquist plot for YbTaO4 EIS device performed under the accumulation region was larger than those under the depletion and inversion regions. Furthermore, the bulk resistance gradually decreased with increasing the RTA temperature, while it clearly increased with increasing the pH value. The YbTaO4-based EIS sensor annealed at 800 °C is suitable for use in future medical and industrial biosensing applications.The effect of RTA treatment on the structural, sensing and impedance characteristics of YbTaO


However, deconvolving the effects of terrestrial contamination from the indigenous hydrogen isotopic compositions of these extraterrestrial materials is not trivial, because chondrites and some achondrites show only small deviations from terrestrial values such that even minor contamination can mask the indigenous values. Here we assess the effects of terrestrial weathering and contamination on the hydrogen isotope ratios and H2O contents of meteoritic minerals through monitored terrestrial weathering of Tissint, a recent Martian fall. Our findings reveal the rapidity with which this weathering affects nominally anhydrous phases in extraterrestrial materials, which illustrates the necessity of sampling the interiors of even relatively fresh meteorite falls and underlines the importance of sample return missions.Determining the hydrogen isotopic compositions and H The potential sources of water for bodies in the inner Solar System include the protosolar nebula, carbonaceous chondrites, and comets. Each of these sources have distinct D/H ratios, expressed as δD values (where δD = [(D/Hsample)/(D/HVSMOW)−1] × 1000 and D/HVSMOW = 155.76 × 10−6). Thus, differences in δD between distinct planetary reservoirs could imply different water sources or secondary processes. For instance, the somewhat lower δD values recently reported in some terrestrial basalts thought to originate from the deep mantle compared to typical mid-ocean ridge basalts has reignited a long-standing controversy about the origin of water on Earth3. As another example, multiple hydrogen isotope investigations of the Martian meteorites have shown δD values ranging from ~−200‰ to ~ + 6000‰ in primary igneous phases in these samples9. While the highest δD values in these Martian meteorite phases are generally thought to represent exchange with crustal fluids that equilibrated with a D-enriched atmosphere on Mars, the lowest have been argued by some authors to represent the Martian mantle9, implying a common origin for the source of water on the Earth and Mars. However, since only ~3% of Martian meteorites are falls and most of the others have had long residence times in the terrestrial environment, it has also been suggested that these low δD Martian values may be a consequence of terrestrial alteration10. Some bulk chondrites, as well as some lunar samples and HED meteorites, have δD values that are apparently similar to terrestrial values15. In such cases in particular, terrestrial contamination makes it difficult to resolve small differences that may exist between an indigenous meteoritic δD value and the terrestrial value.Hydrogen isotope compositions and H16. Furthermore, the degree of this weathering recorded in a meteorite is a function of its terrestrial residence time, climate, soil composition at the recovery site, as well as the meteorite’s physical characteristics and chemical composition17. Weathering can cause visible changes such as rusting of Fe-Ni metal19 and more subtle effects including H2O uptake and isotopic exchange20. Al-Kathiri et al.16 showed a correlation between H2O contents and terrestrial ages of ordinary chondrites, with rapid initial increase of H2O content within 20 ka of terrestrial residence. The Holbrook L6 ordinary chondrite, which fell in 1912, shows a significant shift in δ18O values in samples recovered after a period of 99 years compared to those recovered immediately after the fall21.The lack of detailed characterization of the effects of terrestrial alteration is a major challenge for determining the indigenous δD values of meteorites and those of their corresponding source reservoirs on their parent bodies. Such characterization can be difficult since weathering and alteration in the terrestrial environment usually involves a complex interplay of a variety of factors. Specifically, terrestrial weathering is caused by a wide range of mechanical, chemical and biological processes23. Moreover, previous experimental studies have suggested that contamination and/or exchange of hydrogen in meteorites during residence in a terrestrial environment is minor. For example, the D/H ratio of the Orgueil carbonaceous chondrite did not change significantly after it was immersed in heavy water at room temperature for 30 days24. Robert and Deloule25 studied the hydrogen isotope composition of the Semarkona ordinary chondrite and concluded that “…in light of the large D/H variations observed in Semarkona, the problem of the terrestrial contamination can be ignored.” However, these interpretations have typically been based on analyses of bulk samples or minerals with relatively high H2O concentrations (>0.2 wt.% H2O). In contrast, the relatively low δD values of CR chondrites recovered from the Saharan desert compared to non-Saharan CRs, which have higher bulk δD values, have been attributed to isotopic exchange reactions during hot desert weathering20. Nevertheless, what is not well constrained is the time frame within which H2O abundances and δD values in components of fresh meteorite falls are affected by exposure to the terrestrial environment.In the case of hydrogen isotope compositions, some previous studies have suggested that the effects of terrestrial contamination are negligible for extraterrestrial samples that exhibit δD values far from the terrestrial value. For example, terrestrial contamination has been suggested to be minimal in lunar apatites that have δD values up to 1000‰2O abundances in nominally anhydrous silicate phases with relatively low indigenous H2O contents  (e.g.27), and these phases have been the focus of some recent studies of meteoritic materials. For example, several recent investigations of the Martian meteorites have focused on silicate phases that are likely to record early magmatic conditions prior to significant alteration of δD-H2O systematics by processes such as degassing and fractional crystallization (e.g.9). As such, it is particularly important to characterize the rate at, and degree to which, the δD-H2O systematics in such phases are altered by terrestrial exposure.Finally, advances in analytical techniques and sample preparation protocols have made it possible to investigate hydrogen isotopes and H20. Specifically, if the indigenous δD value of a meteoritic component differs significantly from that of terrestrial reservoirs, then hydrogen contamination will be evident as measured δD values would plot along a mixing trend between the indigenous and terrestrial end-members. It was previously suggested that distinction between secondary aqueous alteration occurring under Martian versus terrestrial conditions can be made based on the measured hydrogen isotope compositions of secondary phases in the Martian meteorites10. In fact, as mentioned earlier, even some primary igneous minerals in the shergottites and nakhlites, which may have interacted and thus exchanged hydrogen with near-surface water reservoirs on Mars, show significant D-enrichment relative to terrestrial reservoirs28. Thus, measuring changes in the δD values of individual phases, such as olivines, maskelynites, and merrillites, in fresh Martian meteorites as a function of the terrestrial residence time of these meteorites can provide a measure of the rate of hydrogen isotopic exchange and addition of terrestrial H2O. Therefore, we report here the results of a terrestrial weathering experiment using the most recent Martian meteorite fall, Tissint29.In meteoritic materials, δD can be used as a fingerprint for terrestrial contamination assuming that there is a measurable difference between the hydrogen isotope compositions of the extraterrestrial sample and the contaminant2O contents of individual phases  in Tissint resulting from different residence times in a desert environment. Fresh samples of Tissint were collected from the dry desert of southern Morocco in October 2011 , prior to any rainfall events. A 1.85 g interior piece of this freshly collected material of Tissint, acquired by the Center for Meteorite Studies, was dry cut into three pieces designated as T0 (0.43 g), T1 (0.70 g), and T3 (0.72 g). One surface on each of these pieces was ground flat and this flattened surface was polished using the dry sample preparation techniques described in9. The T0 sample studied here is the same as the anhydrously prepared thick section (“ATS”) of9; these authors recently reported the results of hydrogen isotope analyses of individual phases in this section. The T0 sample was recovered almost immediately from the Moroccan desert following its fall and is therefore assumed to have experienced minimal, if any, terrestrial alteration. The T1 and T3 samples were placed (after dry polishing one surface of each) in the Sonoran desert  at Arizona State University (ASU). This technique allows high precision δD measurements at high spatial resolution, which was necessary for evaluating the spatial scale of alteration of the hydrogen isotope systematics in the Tissint samples studied here. The hydrogen measured in the SIMS technique is a mixture of signals that includes the indigenous H from within the sample, as well as varying amounts of H from the vacuum chamber within the SIMS instrument  and from contamination on the sample surface31. Several studies have established protocols for sample preparation and SIMS analytical parameters that were utilized in this investigation and that significantly reduced the hydrogen background in the instrument and contamination of the sample surface32. Olivines, maskelynites and merrillites on the dry polished surfaces of the T0 and T1 samples were analyzed. Following its retrieval after 3 years of exposure in the Sonoran desert, the T3 sample was dry cut perpendicular to the exposed polished surface, thereby allowing access to the interior of this sample; olivines were analyzed on the dry polished surface that had been exposed for 3 years (designated as T3′) and in the interior of the T3 sample accessed by the freshly cut surface (designated as T3′′).The D/H ratios (expressed as δD) and H2O contents of maskelynites and merrillites in the polished exposed surface of the T1 sample are similar to those in the minimally altered T0 sample. In contrast, olivines in the polished surfaces of the T0 and T1 samples show marked differences. Specifically, most olivines in the T0 sample have a relatively narrow range of H2O contents (~50–100 ppm), with δD values that vary from ~300 to 600‰; a few olivines have somewhat higher H2O contents (~150 ppm), and these also appear to have systematically lower δD values (0–100‰). In olivines in the polished exposed surface of T1, the lowest H2O contents (~100–150 ppm) are similar to the highest ones in olivines of T0; the δD values in these olivines range from 0 to 200‰. However, the highest H2O contents in these olivines approach 400 ppm (significantly higher than in T0 olivines) and δD values go down to −150‰ . The ranges in H2O contents and δD values of olivines in the polished surface of the T3 sample exposed for 3 years (T3′) are similar to those in olivines of T1 9, this glassy unfractured phase may be less prone to terrestrial contamination than fractured phases like olivine in Tissint  δD values than in T0, but similar ranges of H2O contents and δD as T3′ olivines indicates that, in a desert environment, alteration of H2O-δD systematics in a phase susceptible to weathering  occurs rapidly (within ~1 year) and then reaches a steady state such that longer exposure (up to 3 years) does not cause a significantly greater degree of alteration.The results of our analyses are presented in Fig. int Fig. . The sim2O-δD systematics by analyzing olivines in the interior of the T3 sample   and a range in δD (~100–400‰). In contrast, olivines located ≤2 mm of the exposed surface  have, on average significantly higher H2O contents and lower and more terrestrial-like δD values  still remains relatively pristine.We further investigated the effects of desert weathering on H2O-δD systematics in meteoritic minerals with low indigenous H2O contents located within millimeters of exposed surfaces, particularly if these minerals are also fractured due to processes such as shock. Nevertheless, our results also suggest that such alteration likely reaches a steady state within the first year of exposure, and minerals in the sample interior remain largely unaffected after 3 years of exposure . Given that the duration of our experiment was limited to 3 years, we cannot assess the effects of terrestrial exposure in a desert environment beyond this time period.Therefore, the results reported here clearly demonstrate the rapidity of the effects of terrestrial alteration in a desert environment on H2O content, and their physical properties, such as the presence of fractures), and depend on the duration and conditions of terrestrial exposure. Our results highlight the ease and rapidity with which H2O contents and δD can be affected in minerals with low indigenous H2O content, such as olivine, exposed to a semi-arid desert environment.Terrestrial weathering by aqueous fluids alters the chemical and isotopic properties of meteorites. However, the effects of such weathering are specific to different phases  must be analyzed to ensure the robustness of the δD-H2O data obtained from such samples. We conclude that observed meteorite falls, with subsequent rapid recovery, are best for investigations of H2O-δD systematics in a range of Solar System reservoirs represented by various meteorite types. Moreover, this study underlines the importance of sample return missions, such as OSIRIS-REx and Mars 2020, and appropriate sample curation for determining the indigenous abundances and D/H compositions of water in extraterrestrial materials.The results from this study emphasize the importance of sample selection and preparation in investigations of δD-H9 for preparing a flat, polished surface on each of the T0, T1 and T3 samples; the T0 sample is the same as the anhydrously prepared thick section (“ATS”) described in9. Following emplacement and subsequent retrieval from the Sonoran desert, the T1 and T3 samples were gently brushed with a clean brush with nylon bristles to remove any dirt particles adhering to their exterior surfaces. The T3 sample was then dry cut perpendicular to its polished flat surface; this freshly cut surface was also dry polished as described above. The polished surface of the T3 sample that was exposed to the desert environment (for 3 years) is designated as T3′, while the surface that was cut perpendicular to it  is designated as T3″. Backscattered electron images of each of the three Tissint sections were obtained with the Cameca SX-100 electron microprobe at the University of Arizona. Each of these polished sections of Tissint was then mounted in indium along with individual grains of the standards shown in Table 2O concentrations.We utilized the dry sample polishing technique of2O concentrations were performed on the Cameca IMS-6f at Arizona State University (ASU) using analytical protocols similar to those described in9. For each measurement, a Cs+ primary beam (~10 nA) was rastered over a 30 × 30 μm2 surface area. A field aperture set the analyzed area to 15 μm diameter, which reduced the background associated with the crater edges. Each measurement was comprised of 50 cycles of measuring H− and D− measurements with counting times of 1 s and 10 s, respectively. At the end of each measurement, 16O− was measured. An electron gun was used to maintain charge balance. Vacuum in the analysis chamber was kept at ∼5–10 × 10−10 Torr. The H2O contents were estimated using a H−/16O− vs. H2O calibration curves using several basaltic glasses  from the Indian ridge35, pyroxenes KBH-136 and PMR-5337, as well as and the Durango apatite38 following the method described in9 to account for matrix effects27. The calibration line was corrected for the H2O background and forced through the origin. The H2O contents and δD values of the standards are presented in Table 37 were used to correct for background contamination. The H2O background was estimated to be ~37 ± 1, 50 ± 3 and ~22 ± 1 ppm (2σSD) during the SIMS analytical sessions for the T0, T1 and T3 samples, respectively. The reason for these relatively high background values compared to state-of-the-art SIMS analyses of H2O contents is that the samples investigated in this study were relatively large rock sections with cracks and fractures which did not degas as effectively as the indium-mounted smaller polished fragments or single grains that are typically used for such analyses. These background values are calculated based on the average and the standard deviation (2σSD) of 2–3 analyses of the dry PMR-53 at the beginning of each session, using the H−/16O− vs. H2O calibration line described previously. These background values were subtracted from the H2O concentrations estimated in the phases analyzed in each of these Tissint samples. Errors estimated on H2O concentrations take into account the errors from counting statistics, errors on background estimation, as well as the reproducibility of standards measured during the different analytical sessions over the course of this study. As a result, the external reproducibility (2σSD) of our H2O measurements is estimated to be ± 20%. The δD values are corrected for the instrumental mass fractionation (IMF) and for the background. The δD value for the background was determined on the nominally anhydrous San Carlos olivine and ranged from −150 to −250‰ during different analytical sessions over the course of this study. Errors estimated on δD values take into consideration the error based on counting statistics, as well as the errors in the IMF and on the background δD value.Secondary-ion mass spectrometry (SIMS) measurements of D/H ratios and HSupplementary information


In vivo1H and 31P magnetic resonance spectroscopic imaging (MRSI) provide complementary information on the biology of prostate cancer. In this work we demonstrate the feasibility of performing multiparametric imaging (mpMRI) and 1H and 31P spectroscopic imaging of the prostate using a 31P and 1H endorectal radiofrequency coil (ERC) in combination with a multitransmit body array at 7 Tesla (T).31P transceiver loop coil and 1H receive (Rx) asymmetric microstrip (31P/1H ERC) was designed, constructed and tested in combination with an external 8‐channel 1H transceiver body array coil (8CH). Electromagnetic field simulations and measurements and in vivo temperature measurements of the ERC were performed for safety validation. In addition, the signal‐to‐noise (SNR) benefit of the 1H microstrip with respect to the 8CH was evaluated. Finally, the feasibility of the setup was tested in one volunteer and three patients with prostate cancer by performing T2‐weighted and diffusion‐weighted imaging in combination with 1H and 31P spectroscopic imaging.An ERC with a 31P loop coil showed no differences in the E‐ and B‐fields of the 31P/1H ERC compared with a previously safety validated ERC without 1H microstrip. The hotspot of the specific absorption rate (SAR) at the feed point of the 31P/1H ERC loop coil was 9.42 W/kg when transmitting on 31P at 1 W. Additional in vivo measurements showed a maximum temperature increase at the SAR hotspot of 0.7°C over 6 min on 31P at 1.9 W transmit (Tx) power, indicating safe maximum power levels. When transmitting with the external 1H body array at 40W for 2:30 min, the temperature increase around the ERC was < 0.3°C. Up to 3.5 cm into the prostate the 1H microstrip of the ERC provided higher SNR than the 8CH. The total coil combination allowed acquisition of an mpMRI protocol and the assessment of 31P and 1H metabolites of the prostate in all test subjects.Electromagnetic field simulations of the 31P transceiver and 1H Rx endorectal coil in combination with an 8‐channel transceiver external body array coil and demonstrated its safety and feasibility for obtaining multiparametric imaging and 1H and 31P MRSI at 7T in patients with prostate cancer within one MR examination.We developed a setup with a  Ideally, this would even be combined with T2‐weighted (T2W) and diffusion‐weighted imaging (DWI) of the prostate, as these are paramount in accurate localization and characterization of potential prostate cancer lesions.Proton magnetic resonance spectroscopic imaging (MRSI) is used to study the 1H and 31P spectroscopy and 1H imaging requires a radiofrequency (RF) coil setup with transmit (Tx) and receive (Rx) capabilities on both nuclei,The combination of 31P compared to 1H and the low concentration of 31P metabolites in the human body, 31P spectroscopy of the prostate is challenging. 31P MRSI should preferably be performed at 7 Tesla with an endorectal radiofrequency coil (ERC) to optimize signal‐to‐noise (SNR) and to increase spectral resolution. However, for T2W imaging and DWI of the prostate at 7T, only using an endorectal coil is inadequate. Transmitting radiofrequency pulses with a small local surface coil results in substantial transition bands in T2W and DWI images due to its severely inhomogeneous transmit profile.1+ shimming to provide a homogeneous transmit field.31P, this ERC can also be equipped with 1H receive capabilities to enhance 1H imaging and spectroscopy of the prostate, increasing SNR for 1H spectroscopy and spatial resolution for T2W imaging and DWI. Moreover, the 1H transmit capabilities of the external body array allow the use of the nuclear Overhauser effect for an extra increase in 31P SNR.Because of the lower intrinsic magnetic resonance (MR) sensitivity of 31P Tx/Rx – 1H Rx ERC in combination with an 8‐channel external multitransmit 1H array to enable multiparametric imaging and 1H and 31P spectroscopic imaging of the prostate at 7T. We perform electromagnetic field simulations, phantom measurements and in vivo temperature measurements to validate the safety of the setup. In addition, we evaluate its SNR performance and demonstrate the feasibility of the complete setup in three patients and a volunteer.Therefore, we introduce and evaluate a coil setup with a 22.1One healthy volunteer  and three patients  with histopathologically proven prostate cancer were measured using the setup described below. An intramuscular injection of butylscopolamine bromide  was used to suppress peristalsis. For all volunteer and patient measurements, informed consent was signed and the study was approved by the institutional review board.2.21H and 31P measurements was performed with a custom build SAR supervision system, which measured the time‐averaged input power over 10 s and over 6 min. This input power was not to exceed the safety levels based on local SAR restrictions, otherwise the safety system would stop the measurement.All MR measurements were performed on a 7‐Tesla whole‐body MRI system . Specific absorption rate (SAR) monitoring of the 2.2.11H receive (Rx) coil  were disassembled and equipped with a 1H receive element and a 31P transceiver loop coil. The 1H Rx element was placed directly on the plastic rod1‐ profile was oriented in the same direction as the receive profile of the 31P loop coil. The coil was tuned and matched to 297.2 MHz and a series PIN diode was used for detuning the microstrip during transmit with the external 1H coil. The loop coil was tuned and matched to 50 Ω at 120.3 MHz for 31P at 7T and was mounted on top of the inner balloon . The received signals from the 1H microstrip were routed through a separate coaxial cable to its own interface with integrated 1H preamplifier . These electronics were used to interface the elements in the ERC to the MR system.The mechanical housing and conductors of a 3T endorectal 2.2.21H external multitransmit RF body coil consisted of two arrays containing four microstrip elements with meanders.The 2.2.3in vivo performance of the asymmetric microstrip 1H receive element, we performed in vivo SNR measurements within the prostate to compare the SNR of the ERC with the SNR of the 8‐channel external body array (8CH). Gradient echo (GRE) imaging in transversal and sagittal orientation  = 261 × 380 mm2, matrix = 176 × 256, 3 slices, one average, bandwidth 241 Hz/pixel) of the healthy volunteer was performed with the ERC in situ, using the external body array for transmit and either only the 8‐channel external body array or only the ERC for receive. In the 8CH measurements, the images of the individual coil channels were added using the sum of squares method, after normalizing the individual images to their noise level which was determined within a region of interest (ROI) of air. For both measurements exactly the same measurement parameters were used. The SNR in pixel  of the ERC  and 8CH  measurements was determined by:σERC8CH is an estimate for the noise within the prostate. A region of air was used for this estimation to reduce the influence of physiological noise and variation. σERC8CH was determined by correcting the standard deviation within the air ROI for the Rayleigh distribution in single channel ERC imaging and noncentral chi distribution in sum‐of‐squares multichannel 8CH imaging.To evaluate the The ROI placement for noise calculation was the same for both measurements. The ratio of the SNR of the ERC and the external body array was determined by dividing the respective SNRs:The SNR ratio image was filtered by convoluting the image with a 3 × 3 voxel wide block function.2.331P‐only ERC in combination with the same external body array as used in this study. Here we added a 1H Rx‐only microstrip to their 31P ERC design, which could influence the 31P loop coil performance. Because of the close proximity of the ERC to tissue, this could introduce safety concerns with respect to the maximum allowed 31P transmit power. To demonstrate the safety of the 31P/1H ERC, we performed electromagnetic field simulations of an experimental 31P/1H ERC design which we compared with a design similar to that of Kobus et al.31P/1H ERC to validate the simulation results. The safety characteristics of the 31P/1H ERC were further investigated by performing SAR simulations in a more realistic prostate model and by performing in vivo temperature measurements.Before performing any patient measurements, the setup was thoroughly validated for safety. The SAR safety levels of the setup were in part based on the simulations and measurements of a previously validated design by Kobus et al.Additional phantom measurements using a network analyzer to assess coupling between the ERC and the external coil array are presented in the Supporting Information 2.3.131P loop coil was replaced by a stiff printed circuit board (PCB) plate with the same dimensions, in both simulations and measurements  simulations  at 120.3 MHz of a three‐dimensional (3D) model  of the 31P/1H endorectal coil and a similar ERC without the 1H channel (31P‐only ERC) resembling the design by Kobus et al.r = 79 and a conductivity of σ = 0.47 S/m at a frequency of 120.3 MHz. Additional measurements were performed of the 31P/1H ERC using a setup with field probes (Schmid & Partner Engineering AG (SPEAG), Zürich, Switzerland, probes H3DV7 & ES3DV2) to collect E‐ and B‐field measurements in the sagittal plane in a phantom filled with liquid . To stay within the calibration range of the probes, two different power settings were used: 0.01 W for the B‐field and 0.05 W for the E‐field measurements. The measurements were performed at 120 MHz in a 2D plane using a sampling density of one sample per 5 × 5 mm2 area. A sagittal plane of 150 × 30 mm2 was sampled in the midline of the coil, 10–40 mm above the conductors.To promote reproducibility, the actual 2.3.231P loop coil at 120.3 MHz in a more realistic prostate model were performed to determine the SAR levels at 1 W transmit power. The 31P/1H ERC was compared to the 31P‐only ERC in a model incorporating the different surrounding tissues  of the ues Fig. . Both we2.3.3In vivo temperature measurements were performed with the complete setup: the 31P/1H ERC in combination with the external 1H body array. Three thermocouples were attached to the ERC at different locations . After a localizer and B0 shimming, B1+ shimming was performed by using the B1TIAMO technique 1+ maps and by maximizing the phase coherence of the individual coil channels in an ROI within the prostate. 1H T2‐weighted imaging and diffusion‐weighted imaging were then performed with an in‐plane spatial resolution of respectively 0.43 × 0.43 mm2 and 1.75 × 1.75 mm2.To demonstrate the feasibility of the complete setup, all patient measurements were performed with the external body array coil for 31P MRSI was performed using a 3D phase‐encoded pulse‐acquire sequence with a nonselective BIR‐4 excitation pulse31P SNR.31P MRSI data was fitted using Metabolite Report . PE, PC, GPE, and GPC were modeled as a triplet of three Gaussian peaks with a separation equal to the J‐coupling (6–7.1 Hz) and amplitudes in a 1:2:1 ratio. The PCr lineshape, which was fitted with a single Gaussian shape, was used as a constraint to the linewidth for all signals.31P and 150 ms for 1H) was applied and the spectra were zero‐filled to 2048 points. In the 31P spectra the PCr signal was referenced to 0.0 ppm and in the 1H spectra water was referenced to 4.7 ppm.Proton MRSI was performed using a PRESS‐like sequence with RF refocusing pulses that are both spectrally and spatially selective, such that the signal of a volume of interest (VOI) is received only from the spectral region of interest (2.3–3.3 ppm).331P/1H ERC was constructed according to the circuit and model specifications , MR‐TRUS guided biopsy (one patient) or TRUS biopsy and 3T clinical MRI results (one patient). The ERC’s sensitivity profile was wide enough to perform T2W and DW imaging of the seminal vesicles, apex of the prostate, and anterior regions of the prostate  large prostates, as the sensitivity profile of an ERC drops significantly at large distances.Kobus et al31P spectroscopic imaging was obtained with high quality spectra in all patients, using a spectroscopic imaging method to accurately localize signals. Although the true voxel size was 4.9 or 9.1 cc, it allows voxel localization with respect to the T2W and DWI images within or covering suspicious lesions, instead of relying on coil profile‐based localization with an unlocalized pulse sequence. Using a 31P/1H ERC without external multitransmit array coil also enables the acquisition of 31P and 1H MRSI. However, one would need to revert to the use of many adiabatic RF refocusing pulses and T2‐weighted MRI and DWI becomes particularly problematic with the use of only a small endorectal coil Because of the increased SNR due to the use of 7T MRI in combination with the nuclear Overhauser effect, 1H MRSI was also obtained with a high SNR and with well distinguishable tCho and Spm signals, but it suffered from lipid contamination in some cases. This is most likely caused by B0 inhomogeneities, the small chemical shift separation between the citrate and lipid signals and the spectral selectivity of the spectral‐spatial pulses of the 1H spectroscopy sequence. 1H MRSI was performed with a corrected voxel size of 1.4 cc, enabling voxel localization within or covering lesions. The spectral spatial refocusing pulses selectively inverted the 3.1 ppm resonance of Spm, refocusing its J‐coupling at the echo time,In the seminal vesicles, an increased PC/PE ratio and a high tCho signal with an almost absent Spm peak were found .1H spectroscopy tCho/Spm could be used, as tCho tends to be elevated in cancer lesions, whereas spermine tends to decrease in cancer tissue.Although there is a correlation between 531P Tx/Rx 1H Rx endorectal coil in combination with an external multitransmit 1H body array enables high resolution multiparametric MR imaging and 1H and 31P spectroscopy of the prostate at 7T. The setup does not pose any safety issues and provides superior 1H SNR locally in the prostate in comparison with a 1H external body array alone. The clinical data shown in this study are anecdotal but they illustrate how the proposed setup would be particularly suitable to combine anatomically and functionally with metabolic imaging in prostate cancer.Using a The authors have no relevant conflicts of interest to disclose.Figure S‐1: B1− simulations of the 1H Rx asymmetric microstrip  and a loop coil with similar length .Figure S‐2: A schematic transversal overview of the complete in vivo measurement setup, including a transversal flip angle map through the abdomen  when transmitting with the combined 8‐channel coil array after B1+ shimming.Figure S‐3: Transversal T2W imaging of a patient  in the seminal vesicles (a), the mid‐prostate (b) and the apex (c) of the prostate.Figure S‐4: ADC map (a), T2W image (b) and 31P MRSI spectrum (c) of a patient  with prostate cancer (Gleason 3+4).Table S‐1: Coupling between 31P/1H endorectal coil and external 1H body array in dB.Click here for additional data file.


Scientific Reports 10.1038/srep32062, published online 31 August 2016Correction to: This Article contains errors.In Figure 3C, the merge images of confocal micrographs for Apo-bLf 10 nM and Dox 1.5 µM are incorrect and represents a different replicate of same treatment group.Due to an error that occurred at the time of figure assembly, the representative image for 48 h Fe-bLf sample in Figure 5F was included as a duplicate image of 96 h Apo-bLf sample. In addition, for 48 h untreated sample, the representative image is incorrect and shows a different sample.The correct versions of Figures 3C and 5F are given below as Figures This change does not affect the conclusions of the Article. The authors apologize for the errors.


DEFB1, OMIM#602056) with the risk of developing type 2 diabetes mellitus (T2DM) in a group of Mexican patients.The aim of this study was to establish the association of two polymorphisms of the β‐defensin 1 gene . In addition, the linkage disequilibrium analysis showed that AC haplotype was associated with an increased risk of developing T2DM . The in‐silico analysis showed that the −44 C allele produces a binding site for the transcription factor Ikaros (IK).Under dominant and additive models adjusted for the risk factors, the C allele of the −44 C/G polymorphism was associated with increased risk of T2DM (OR = 1.63, 95% CI = 1.07–2.48, pCDEFB1 gene could increase the migration of the macrophages to pancreatic islets accelerate the β‐cell dysfunction in T2DM.This study demonstrates that the C allele of −44 C/G polymorphism, as well as haplotype AC are associated with the presence of T2DM in the Mexican population. The variation in this polymorphism of the  Blood pressure was measured three times by trained personnel using sphygmomanometry. Hypertension was defined as systolic blood pressure ≥140 mmHg and/or diastolic blood pressure ≥90 mmHg or the use of oral antihypertensive therapy. Plasma lipid concentrations were determined after a 12‐hr overnight fast, EDTA blood samples were drawn and centrifuged within 15 minutes after collection; plasma was separated into aliquots and were analyzed within 24‐hr after blood sample collection. Cholesterol and triglycerides plasma concentrations were determined by enzymatic/colorimetric assays . The phosphotungstic acid‐Mg2+ method was used to determine HDL‐C concentrations. LDL‐cholesterol (LDL‐C) was estimated in samples with a triglyceride level lower than 400 mg/dl, using the Friedewald formula, modified by DeLong . Individuals with type 1 diabetes (T1D), maturity‐onset diabetes of the young (MODY), secondary diabetes due to hemochromatosis, cystic fibrosis, or pancreatitis or individuals with a family member with T1D were excluded from the study. The 522 healthy individuals  belong from the Genetics of Atherosclerosis Disease (GEA) Mexican study. All subjects were asymptomatic and apparently healthy individuals without a family history of premature CAD, atherosclerosis, and T2DM who were recruited during June 2009 to June 2013 among blood bank donors and with the assistance of brochures posted in social service centers. The exclusion criteria included the use of anti‐dyslipidemic or anti‐inflammatory drugs at the time of the study, congestive heart failure, and liver, renal, thyroid, or oncological disease  and 552 healthy controls (26–78 years old). From July 2010 to July 2015, 252 patients  were referred to the Instituto Nacional de Cardiologia Ignacio Chavez. The diagnosis of T2DM was made according to the American Diabetes Association (ADA) and American Heart Association (AHA) based on three criteria, (a) an HbA1c value ≥6.5%, (b) fasting plasma glucose levels ≥126 mg/dl, (c) 2‐hr plasma glucose ≥200 mg/dl during an oral glucose tolerance test. The test was performed as described by the World Health Organization, using a glucose load containing the equivalent of 75 g anhydrous glucose dissolved in water  and −44 C/G  polymorphisms was made using 5′ exonuclease TaqMan assays as indicated by the manufacturer . Samples previously sequenced were included as positive controlsThe GenBank reference for 2.3https://snpinfo.niehs.nih.gov/cgi-bin/snpinfo/snpfunc.cgi) program. This program analyze the location of the SNP  and its possible functional effects, such as amino acid changes in protein structure, transcription factor binding sites in promoter or intronic enhancer regions, and alternative splicing regulation by disrupting exonic splicing enhancers or silencers  were equal or >10%. When a principal effect model was reached, the effect modification was also tested and interaction terms were constructed between the polymorphisms and various variables; the terms were included in the model when the significance of the p‐value was higher or equal to 0.05. All p‐values were corrected (pC) by the Bonferroni test. pC values <0.05 were considered statistically significant, and all odds ratios (OR) are presented with 95% confidence intervals. The occurrence of the T2DM in our population was based in the OR values: OR = 1 does not affect odds of developing T2DM, OR > 1 is associated with higher odds of developing T2DM, and OR < 1 is associated with lower odds of developing T2DM. The linkage disequilibrium analysis  of the polymorphisms as well as the haplotype construction was performed with Haploview version 4.1 . The analysis of data was performed with SPSS version 18.0  statistical package. The statistical power to detect association of the polymorphisms with T2DM was 0.80 and was estimated with QUANTO software (https://biostats.usc.edu/software).Considering that the continuous variables such as age, body mass index, blood pressure, glucose, total cholesterol, HDL‐C, LDL‐C, and triglycerides did not have normal behavior, they were analyzed using the Mann–Whitney 33.1Baseline characteristics of the T2DM patients and healthy controls included in the study are presented in Table 3.2DEFB1 gene were in Hardy–Weinberg equilibrium. The 5′UTR −20 G/A (rs11362) polymorphism frequency was similar in patients with T2DM and healthy controls. However, under dominant and additive models adjusted for gender, age, BMI, hypertension, dyslipidemia, and alcohol consumption, the C allele of the 5’UTR −44 C/G (rs1800972) polymorphism was associated with increased risk of developing T2DM  (Table The observed and expected frequencies in the two polymorphic sites of the 3.3DEFB1 5′UTR −20 G/A [rs11362] and DEFB1 5′UTR −44 C/G [rs1800972]) showed strong linkage disequilibrium (D′ > 0.94) and four haplotypes were constructed: H1 (AG), H2 (GC), H3 (GG), and H4 (AC). Haplotype H4 was associated with an increased risk of developing T2DM as compared to controls , whereas the distribution of the haplotypes H1, H2, and H3 was similar in T2DM patients and healthy controls  polymorphism produces a binding motif for the breast cancer transcription factor (BRCA). The analysis also revealed that the C allele of the DEFB1 5′UTR −44 C/G (rs1800972) polymorphism generates binding motifs for ikaros transcription factor (IK).The functional prediction analysis showed that the presence of the 4DEFB1 gene were analyzed in T2DM patients and healthy controls. The distribution of 5′UTR −20 G/A (rs11362) was similar in both groups. Nonetheless, the analysis of the 5′UTR −44 C/G (rs1800972) polymorphism showed that the C allele was associated with an increased risk of developing T2DM under dominant and additive model. To the best of our knowledge, this is one of the few studies that describe the association of these polymorphisms with T2DM. Agreement with our results, Nemeth et al., report that the −44 CC genotype of the 5′UTR −44 C/G polymorphism was associated with increased risk for developing diabetes (OR = 2.05) in a Hungarian population. In addition, the authors reported that the frequency of the −44 GG genotype was lower in patients with diabetes (2%) than in the healthy controls (9%), suggesting that this genotype has a protective role in the development of diabetes . As can be seen, this haplotype bears the C allele, that was associated with the disease in the independent analysis of the polymorphisms. This result corroborates the participation of this allele as a risk factor in the developing of the T2DM. Nonetheless, we considered that our findings, as well as the reported by Nemeth et al.  located in the 5′ untranslated region (UTR) of h et al. , need toDEFB1 5′UTR‐20 G/A and DEFB1 5′UTR‐44 C/G polymorphisms. Although the 5′UTR‐20 G/A SNP was not associated with T2DM, the analysis in silico showed that the A allele produces a binding motif for the BRCA1 transcription factor. The BRCA1 tumor suppressor is a protein that has an important role in multiple biochemical and biological functions such as DNA repair, replication, cell cycle checkpoints, protein ubiquitination, transcriptional regulation, chromatin remodeling, and apoptosis  in human breast cancer cell line (MCF7). In line with this data, Shukla et al. (BRCA1 results in increased expression of IRS‐1 (insulin receptor substrate‐1) and increased levels of IGF‐1R in BRCA1‐deficient mice, and in cultured human cells. On the other hand, our in‐silico analysis showed that the −44 C allele produces a binding site for the Ikaros (IK) transcription factor; IK factor regulates the expression of several other transcription factors as Cdx4 that participates in the activation of the STAT4, and in the regulation of the expression of several pro‐inflammatory cytokines  affect the regulation the insulin‐like growth factor receptor (IGF‐IR), and (b) heighten expression of pro‐inflammatory cytokines. On the other hand, recent studies showed the β‐defensin 1 has an important role as chemoattractant molecule of macrophages through its interaction with CCR6  associated with the presence of T2DM. Nonetheless, due to the number of individuals included in our study and the specific genetic characteristics of the Mexican population, we considered that additional studies in a larger number of individuals and in other populations with different ethnic origins could help to define the true role of this polymorphism as susceptibility marker for developing T2DM and others inflammatory events.In summary, this study demonstrates that the The authors declare that there are no competing interests regarding the publication of this article.Gilberto Vargas‐Alarcon and Marco Antonio Martinez‐Rios contributed equally to this study. Gilberto Vargas‐Alarcon and Jose Manuel Fragoso were responsible for the conception and design of the study. Jose Manuel Rodriguez‐Perez, Gabriel Herrera‐Maya, and Carlos Posadas‐Romero, participated in the generation and collection of the samples. Marco Antonio Martinez‐Ríos, Marco Antonio Peña‐Duque, and Nonanzit Perez‐Hernandez, contributed in the analysis, and interpretation of data. Drafting or revision of the manuscript was handled by Gilberto Vargas‐Alarcon, Oscar‐Perez‐Mendez and Jose Manuel Fragoso.


P = 0.013 and 0.004, respectively; odds ratio 2.529, 95% confidence interval 1.325–4.827). The genotype and allele frequencies of the other 16 SNP sites were not found to be different between the PD group and the control group. rs2550936, rs3776510, and rs429699 were selected to construct the haplotypes; no significant difference was found in a frequency of 5 haplotypes between the PD group and the control group. These results suggest that the SLC6A3 variant in rs40184 A allele may increase the risk of PD in northwest Han population and may be a biomarker of PD.Parkinson's disease (PD) is one of the most common neurodegenerative disorders in neurology. It is possible that multifactorial and genetic factors are related to its pathogenesis. Recently, there have been reports of SLC6A3 genetic variants leading to PD. However, the role of 3′ end of SLC6A3 in PD is less studied in different ethnic groups. To explore the roles of 3′ end of SLC6A3 in PD development, 17 SNP sites in 3′ end of SLC6A3 were analyzed in 360 PD patients and 392 normal controls of Han population residing in northwest of China. The significant difference of gene type and allele frequencies between the PD and control groups was detected only in rs40184 ( Parkinson's disease (PD) is a neurodegenerative disease; its prevalence increases with age, and PD influences 1% of the population over 60 years . Due to Since PD is characterized by the selective loss of dopaminergic neurons, genes that affect the expression of dopaminergic neurons may become candidate genes for PD. The evidence from genetic studies revealed that variants in SLC6A3 were associated with PD –13. The A cohort of 360 Chinese Han PD patients (63.5 ± 10.4 years) were enrolled. All the patients came from the Inner Mongolia Medical University Affiliated Hospital, Hohhot, China, and Bayannaoer City Hospital, Bayannaoer, China, between 2015 and 2018. All patients live in northwest of China and were not related to each other. All patients were examined by experienced neurologists, and the diagnosis of PD was based on clinical criteria . The conhttp://www.hapmap.org), which is based on pairwise r2 ≥ 0.8 and minor allele frequency (MAF) ≥0.1. In this study, we chose 17 tagSNPs  from the 3′ end of SLC6A3.TagSNPs were selected from the Chinese HapMap database , which was stored at −20°C. Gene typing was performed using the polymerase chain reaction (PCR)/ligase detection reaction assay. Primers were synthesized by HAYU Biological Engineering LTD in Shanghai; the information of the primers is shown in μl, which contained 1X PCR buffer, 3.0 mM/L·MgCl2, 2.0 mM/L deoxynucleotide triphosphate, 0.5 μmol/μl primer mix, 5 U/μl Qiagen HotStarTaq Polymerase , 1X Q-solution, and 50 ng/μl genomic DNA. The thermal cycle was carried out in GeneAmp PCR system 9600 . The initial denaturation was 2 min at 95°C, followed by 40 cycles of denaturation at 94°C for 30 s, annealing at 56°C for 90 s, and extension at 72°C for 1 min. The final extension of 72°C is 10 min.The multiplex PCR methods were used to amplify the target DNA sequences. The final volume of PCRs for each subject was 20 μl for each subject, which contains 1X NEB Taq DNA ligase buffer 1 μl, 2 pmol of each probe mix 1 μl, 5 U/μl Taq DNA ligase 0.05 μl , and 4 μl multi-PCR product. A total of 40 ligase detection reaction cycles were performed under conditions of 92°C for 2 min, 94°C for 15 s, and 50°C for 25 s. The fluorescent products of ligase detection reaction were identified by PRISM 3730 (ABI). The experimental methods are similar to Chang et al. [The total volume of the ligase detection reaction is 10 P value of 0.05 was considered to have statistical significance.Statistical analysis was carried out by the Statistical Program for Social Sciences (SPSS version 11.0). Hardy-Weinberg equilibrium of each group was determined by using the chi-squared test. Allele and genotype frequencies between groups were studied using SHEsis software . We use P > 0.05). Statistically significant differences in genotype and allele frequencies were found in the SLC6A3 variant rs40184 between PD  and controls . The frequency of the minor A allele was 29.2% in patients and 14.0% in controls. There was no significant difference in genotype and allele frequencies for other 16 SNP polymorphism sites in the 3′ end of SLC6A3.The genotype and allele frequencies of the 3′ end of SLC6A3 are summarized in LD plots of the SLC6A3 in the study are shown in PD is a complex disease caused by age and environmental and genetic factors. Finding the genetic susceptibility factors for PD may help identify the individuals at risk and design more specific prevention or treatment options for them. SLC6A3 gene encodes DAT, which comprises 15 exons spanning 60 kb on chromosome 5p15.32 . The stuHere, we conducted a case-control study of 752 participants in the northwest Han population to further investigate the role of the 3′ end of SLC6A3 in the development of PD. We found that only the SLC6A3 variant rs40184 had statistically significant differences in genotypes and alleles, which may be related to PD. The minor A allele of rs40184 may lead to an increased risk of PD in northwest Han Chinese, and it may be a marker of PD. However, the other 16 polymorphic sites were not found to be related to PD. The reason why the other 16 sites had negative results may be the insufficient number of samples in this study and that these 16 sites may be rare genotypes or alleles in the Han population. Haplotype analysis is thought to be much more powerful than single-nucleotide polymorphism sites in correlation studies . HaplotyAt present, more than 1,500 genes are known to be closely related to the occurrence of diseases. For a long time, coding areas have been a major research direction of genetic diseases and only a small part of noncoding area has been proved to be a useful component that can help genes to be turned on and off to regulate gene expression. The normal expression of genes cannot be separated from the participation of regulatory elements. Abnormalities in certain regulatory components can also lead to mutations in the corresponding gene-coding regions. Therefore, we can assume that the variant rs40184 in the noncoding area of SLC6A3 may lead to differences in susceptibility to PD in our study.In summary, our findings showed a link between the 3′ end of SLC6A3 gene variant rs40184 and PD in northwest Han population. Given that different populations are genetically heterogeneous and mutations have a specific population frequency, larger sample size studies are needed to confirm the correlation between the 3′ end of SLC6A3 variant and PD in independent larger cohorts and in different geographical origins. Moreover, functional studies of the 3′ end of SLC6A3 need to be carried out to further understand the role of SLC6A3 in PD. Further research is likely to find genetic variations that are risky or protective of PD, which is important for the prevention and treatment of PD.