Low-temperature Electron Paramagnetic Resonance Research Articles

Photodarkening-resistance improvement of Yb3+/Al3+ co-doped silica fibers fabricated via sol-gel method.

Yb3+/Al3+ co-doped silica fibers (YDFs) with almost identical core glass compositions were prepared using the sol-gel and modified chemical vapor deposition (MCVD) methods. The photodarkening (PD) and laser performance before and after the PD process were tested under 974 nm pumping. The doping homogeneity of Yb3+ ions and clusters of Yb3+ ions in preform slices of these two fibers were investigated via optical absorption spectroscopy, photoluminescence emission spectra, electron probe microanalysis (EPMA), and low-temperature (4 K) electron paramagnetic resonance (EPR). It is known that the PD resistance of YDFs prepared via the sol-gel method is significantly better than that of YDFs prepared via MCVD under the same test conditions. EPMA mapping reveals that the doping homogeneity of Yb3+ ions in the sol-gel fiber core glass is better than that in the MCVD fiber. The low-temperature (4 K) EPR and cooperative luminescence spectra of Yb3+ ions indicate that the clustering degree of Yb3+ ions in the sol-gel fiber is lower than that in the MCVD fiber. In the absorption and emission spectra, small amounts of Yb2+ ions are observed in the preform slice from the sol-gel method. A model of the color-center generation in the PD process was proposed to explain the mechanism of PD resistance improvement for the YDFs fabricated via the sol-gel method.

Low-Temperature EPR Spectroscopy as a Probe-Free Technique for Monitoring Oxidants Formed in Tumor Cells and Tissues: Implications in Drug Resistance and OXPHOS-Targeted Therapies.

Oxidants formed from oxidative and nitrative metabolism include reactive oxygen species (ROS) such as superoxide, hydrogen peroxide/lipid hydroperoxides and reactive nitrogen species (RNS) (e.g., peroxynitrite [ONOO-] and nitrogen dioxide), and reactive halogenated species (e.g., hypochlorous acid [HOCl]). Increasingly, ROS and RNS are implicated in tumorigenesis as well as tumor growth, progression, and metastasis. Recently, ROS were implicated in drug resistance, metabolic reprogramming, and T-cell metabolism in immunotherapy. Mostly, fluorescent probes have been used in cell culture systems. The identity of species is obtained by LC-MS analyses of diagnostic marker products. However, extrapolation of these assays to cancer xenografts is difficult if not impossible. Thus, development of a probe-free assay for monitoring and assessing oxidant formation in tumor cells and tumor xenografts is critical and timely. Here, we describe the use of ex vivo electron paramagnetic resonance (EPR) spectroscopy at cryogenic temperatures as a uniquely useful probe-free technique for assessing intracellular oxidation and oxidants via EPR signals from redox centers, particularly iron-sulfur clusters, in mitochondrial and cytosolic redox proteins. Examples of cancer cells subjected to inhibition of mitochondrial oxidative phosphorylation are presented. This ex vivo methodology can be readily extended to monitor oxidant formation in tumor tissues isolated from mice and humans.

Genotoxic Responses of Mitochondrial Oxygen Consumption Rate and Mitochondrial Semiquinone Radicals in Tumor Cells

Our recent report demonstrated that genotoxic stimuli enhance mitochondrial energy metabolism in various tumor cell lines. However, the mitochondrial response against genotoxic stimuli has not been fully elucidated. In this study, to investigate mitochondrial functions in X-irradiated cells, the oxygen consumption rate (OCR) in human cervical adenocarcinoma HeLa cells was examined by electron spin resonance (ESR) spectroscopy with lithium 5,9,14,18,23,27,32,36-octa-n-butoxy-2,3-naphthalocyanine. ESR oximetry demonstrated that basal respiration, ATP-linked respiration, proton leak, maximal respiration, and reserve capacity increased in HeLa cells 24 h after X-irradiation. However, a flow cytometric analysis using MitoTracker Green showed that mitochondrial mass also increased following X-irradiation. When the OCR was standardized to the mitochondria membrane mass, the radiation-induced increases in the respiratory parameters disappeared. This finding indicated that the radiation-induced increase in cellular OCR was explained by an increase in mitochondrial mass but not by the activation of mitochondrial respiratory-related enzymes. In addition, mitochondrial semiquinone radicals at g = 2.004 were detected by low-temperature (110 K) ESR spectroscopy. The ESR signal intensity of semiquinone radicals was enhanced by X-irradiation, suggesting an increase in the electron flow in the electron transport chain. These data will be important to understand the mechanism of radio-sensitization by mitochondria-targeting reagents in tumor cells.

Evaluation of mitochondrial redox status and energy metabolism of X-irradiated HeLa cells by LC/UV, LC/MS/MS and ESR

To evaluate the metabolic responses in tumour cells exposed to ionizing radiation, oxygen consumption rate (OCR), cellular lipid peroxidation, cellular energy status (intracellular nucleotide pool and ATP production), and mitochondrial reactive oxygen species (ROS), semiquinone (SQ), and iron–sulphur (Fe−S) cluster levels were evaluated in human cervical carcinoma HeLa cells at 12 and 24 h after X-irradiation. LC/MS/MS analysis showed that levels of 8-iso PGF2α and 5-iPF2α-VI, lipid peroxidation products of membrane arachidonic acids, were not altered significantly in X-irradiated cells, although mitochondrial ROS levels and OCR significantly increased in the cells at 24 h after irradiation. LC/UV analysis revealed that intracellular AMP, ADP, and ATP levels increased significantly after X-irradiation, but adenylate energy charge (adenylate energy charge (AEC) = [ATP + 0.5 × ADP]/[ATP + ADP + AMP]) remained unchanged after X-irradiation. In low-temperature electron spin resonance (ESR) spectra of HeLa cells, the presence of mitochondrial SQ at g = 2.004 and Fe–S cluster at g = 1.941 was observed and X-irradiation enhanced the signal intensity of SQ but not of the Fe–S cluster. Furthermore, this radiation-induced increase in SQ signal intensity disappeared on treatment with rotenone, which inhibits electron transfer from Fe–S cluster to SQ in complex I. From these results, it was suggested that an increase in OCR and imbalance in SQ and Fe–S cluster levels, which play a critical role in the mitochondrial electron transport chain (ETC), occur after X-irradiation, resulting in an increase in ATP production and ROS leakage from the activated mitochondrial ETC.

Electron Spin Resonance Investigations on Perovskite Solar Cell Materials Deposited on Glass Substrate

ABSRTACTIn this article, we report low-temperature electron spin resonance (ESR) investigations carried out on solution processed three-layer inverted solar cell structures: PC61BM/CH3NH3PbI3/PEDOT:PSS/Glass, where PC61BM and PEDOT:PSS act as electron and hole transport layers, respectively. ESR measurements were conducted on ex-situ light (1 Sun) illuminated samples. We find two distinct ESR spectra. First ESR spectra resembles a typical powder pattern, associated with gx = gy = 4.2; gz = 9.2, found to be originated from Fe3+ extrinsic impurity located in the glass substrate. Second ESR spectra contains a broad (peak-to-peak line width ∼ 10 G) and intense ESR signal appearing at g = 2.008; and a weak, partly overlapped, but much narrower (peak-to-peak line width ∼ 4 G) ESR signal at g = 2.0022. Both sets of ESR spectra degrade in intensity upon light illumination. The latter two signals were found to stem from light-induced silicon dangling bonds and oxygen vacancies, respectively. Our controlled measurements confirm that these centers were generated during UV-ozone treatment of the glass substrate –a necessary step to be performed before PEDOT:PSS is spin coated. This work forms a significant step in understanding the light-induced- as well as extrinsic defects in perovskite solar cell materials.

Magnetic characterization of microcrystalline Na3Ln0.99–xEr0.01Crx(PO4)2 orthophosphates synthesized by Pechini method (Ln = La, Gd)

Abstract Na3Ln(PO4)2 orthophosphates (Ln = La, Gd) doped with Er3+ and co-doped with Cr3+ ions were synthesized by Pechini method and characterized by electron paramagnetic resonance (EPR) and magnetic susceptibility measurements. Low temperature EPR spectra were detected and analyzed in terms of temperature dependence and the structure of the obtained materials. They show that erbium and chromium ions substitute Ln3+ and also Na+ ions or Na+ channels forming complex EPR spectra. Both kinds of ions reveal ferromagnetic type of interaction which shows some anomaly at the temperature between 10 K and 15 K. Magnetic susceptibility reveals a weak antiferromagnetic kind of interaction dominating in the whole temperature range, from 3.5 to 300 K.

Crystal-field effects in Er3+ - and Yb3+ -doped hexagonal NaYF4 nanoparticles

Since the up-conversion phenomenon in rare-earths (REs) doped ${\mathrm{NaYF}}_{4}$ is strongly affected by the crystal electric field (CF), determining the CF parameters, wave functions, and scheme of the energy levels of the RE $J$ multiplets could be crucial to improve and tune the up-conversion efficiency. In this work, the temperature and magnetic field dependent magnetization of ${\mathrm{NaY}}_{1\ensuremath{-}x}{[\mathrm{Er}(\mathrm{Yb})]}_{x}{\mathrm{F}}_{4}$ hexagonal nanoparticles (NPs) is reported. The data were best fit using the appropriated CF Hamiltonian for the $J=15/2\phantom{\rule{0.16em}{0ex}}(J=7/2)$ ground state multiplet of ${\mathrm{Er}}^{3+}\phantom{\rule{0.16em}{0ex}}({\mathrm{Yb}}^{3+}$) ions. The ${B}_{2}^{0}, {B}_{4}^{0}, {B}_{6}^{0}$, and ${B}_{6}^{6}$ CF parameters were considered in the Hamiltonian for RE ions located at the hexagonal ${\mathrm{C}}_{3h}$ point symmetry site of the ${\mathrm{NaYF}}_{4}$ host lattice. These results allowed us to predict an overall CF splitting of $\ensuremath{\sim}214$ ($\ensuremath{\sim}356$ K) for ${\mathrm{Er}}^{3+}$ (${\mathrm{Yb}}^{3+}$) and the wave functions and their energy levels for the $J=15/2$ ($J=7/2$) ground state multiplet which are in good agreement with the low temperature electron spin resonance experiments. Besides, our measurements allowed us to calculate all the excited CF $J$ multiplets that yield to a good estimation of the up-conversion light emission linewidth. The nonlinear optical light emission of the studied ${\mathrm{NaY}}_{1\ensuremath{-}x}{[\mathrm{Er}(\mathrm{Yb})]}_{x}{\mathrm{F}}_{4}$ hexagonal NPs was also compared with the most efficient up-conversion codoped ${\mathrm{NaY}}_{1\ensuremath{-}x\ensuremath{-}y}{\mathrm{Er}}_{x}{\mathrm{Yb}}_{y}{\mathrm{F}}_{4}$ hexagonal NPs.

Electrochemistry and electron paramagnetic resonance spectroscopy of cytochrome c and its heme-disrupted analogs

Cytochrome c (cyt c) is one of the most studied conjugated proteins due to its electron-transfer properties and ability to regulate the processes involved in homeostasis or apoptosis. Here we report an electrochemical strategy for investigating the electroactivity of cyt c and its analogs with a disrupted heme moiety, i.e. apocytochrome c (acyt c) and porphyrin cytochrome c (pcyt c). The electrochemical data are supplemented with low-temperature and spin-probe electron paramagnetic resonance (EPR) spectroscopy. The main contribution of this report is a complex evaluation of cyt c reduction and oxidation at the level of surface-localized amino acid residues and the heme moiety in a single electrochemical scan. The electrochemical pattern of cyt c is substantially different to both analogs acyt c and pcyt c, which could be applicable in further studies on the redox properties and structural stability of cytochromes and other hemeproteins.

(Invited) Probing Dopants in 2H MoS2 Crystals and 2D Layers by Electron Paramagnetic Resonance: Identification and Quantification

Low-temperature electron spin resonance (ESR) study on the atomic nature of doping in geological p-type 2H MoS2 crystals reveals two main types of responses. The major one, of axial symmetry, stems from As acceptors, with As substituting for S, which, occurring with density ~2.4x1017 cm-3, predominantly accounts for the p-type doping. Its ESR spectrum points to manifest non-uniform As dopant distribution, i.e., substantial clustering, with only ~13% of the As response originating from single decoupled As impurities. A second, much weaker so far unknown signal, of spin density ~5x1014 cm-3, shows lower than axial symmetry, likely orthorhombic (C 2v). Based on salient ESR characteristics, it is assigned to the Pb impurity substituting for Mo, the center operating as acceptor. The dopant activation energy, Ea, was assessed through cautious monitoring of the signal intensity vs. temperature. For As, this resulted in Ea = 0.7 ± 0.2 meV, unveiling As as a noticeable shallow acceptor. The adduced insight may contribute to developing tunable, robust p-type doping strategies for few-layer MoS2 application in future nano-electronic devices.

Zn-Assisted TiO2–x Photocatalyst with Efficient Charge Separation for Enhanced Photocatalytic Activities

A new Zn-assisted method has been employed to synthesize reduced TiO2(TiO2–x) photocatalyst via a one-step hydrothermal process. In order to prevent the oxidation of TiO2–x in air, hydrofluoric acid is introduced in the preparative process for the stabilization of the Ti3+ and oxygen vacancies, as confirmed by low-temperature electron paramagnetic resonance. The obtained reduced TiO2 presents a wide-spectrum solar light absorption, including the near-infrared region. In addition, {110}–{111} and {101}–(001) dual-facet exposures are generated by Cl- and F-based surface-terminated reagents, respectively. The generated {001} and {101} facets on reduced TiO2 samples act as hole and electron collectors, respectively, which contributes to the charge separation of the catalyst. Finally, the synergistic effect between Ti3+ doping and dual-facet exposure results in the high photocatalytic performance for degradation of Rhodamine B and formic acid.

INFLUENCE OF EXOGENOUS LACTOFERRIN ON THE OXIDANT/ ANTIOXIDANT BALANCE AND MOLECULAR PROFILE OF HORMONE RECEPTOR-POSITIvE AND -NEGATIvE HUMAN BREAST CANCER CELLS IN VITRO

To investigate the mechanisms of cytotoxic activity and pro-/antioxidant effect of lactoferrin on hormone receptor-positive and receptor-negative breast cancer cells in vitro. The study was performed on receptor-positive (MCF-7, T47D) and receptor-negative (MDA-MB-231, MDA-MB-468) human breast cancer cell lines. Immunocytochemical staining, flow cytometry, low-temperature electron paramagnetic resonance, and the Comet assay were used. Upon treatment with lactoferrin, the increased levels of reactive oxygen species (ROS) (p < 0.05), NO generation rate by inducible NO-synthase (p < 0.05) and the level of "free" iron (p < 0.05) were observed. Moreover, the effects of lactoferrin were more pronounced in receptor-negative MDA-MB-231and MDA-MB-468cells. These changes resulted in increased expression of proapoptotic Bax protein (p<0.05), reduced expression of the antiapoptotic Bcl-2 protein (p < 0.05) and level of not-oxidized mitochondrial cardiolipin (1.4-1.7-fold, p< 0.05). This, in turn, caused an increase in the percentage of apoptotic cells (by 14-24%, p < 0.05). Cytotoxic effects of lactoferrin were accompanied by an increase in the percentage of DNA in the comet tail and blocking cell cycle at G2/M phase, especially in receptor-negative cell lines. The study showed that exogenous lactoferrin causes a violation of an antioxidant balance by increasing the level of ROS, "free" iron and NO generation rate, resalting in the blocking of cell cycle at G2/M-phase and apoptosis of malignant cells.

Modified Metformin as a More Potent Anticancer Drug: Mitochondrial Inhibition, Redox Signaling, Antiproliferative Effects and Future EPR Studies.

Metformin, one of the most widely prescribed antidiabetic drugs in the world, is being repurposed as a potential drug in cancer treatment. Epidemiological studies suggest that metformin exerts anticancer effects in diabetic patients with pancreatic cancer. However, at typical antidiabetic doses the bioavailability of metformin is presumably too low to exert antitumor effects. Thus, more potent analogs of metformin are needed in order to increase its anticancer efficacy. To this end, a new class of mitochondria-targeted metformin analogs (or mito-metformins) containing a positively-charged lipophilic triphenylphosphonium group was synthesized and tested for their antitumor efficacy in pancreatic cancer cells. Results indicate that the lead compound, mito-metformin10, was nearly 1000-fold more potent than metformin in inhibiting mitochondrial complex I activity, inducing reactive oxygen species (superoxide and hydrogen peroxide) that stimulate redox signaling mechanisms, including the activation of adenosinemonophosphate kinase and inhibition of proliferation of pancreatic cancer cells. The potential use of the low-temperature electron paramagnetic resonance technique in assessing the role of mitochondrial complexes including complex I in tumor regression in response to metformin and mito-metformins in the in vivo setting is discussed.

Synthesis and Characterization of 5- and 6- Coordinated Alkali Pertechnetates

ABSTRACTThe local chemistry of technetium-99 (99Tc) in oxide glasses is important for understanding the incorporation and long-term release of Tc from nuclear waste glasses, both those for legacy defense wastes and fuel reprocessing wastes. Tc preferably forms Tc(VII), Tc(IV), or Tc(0) in glass, depending on the level of reduction of the melt. Tc(VII) in oxide glasses is normally assumed to be isolated pertechnetate TcO4-anions surrounded by alkali, but can occasionally precipitate as alkali pertechnetate salts such as KTcO4and NaTcO4when Tc concentration is high. In these cases, Tc(VII) is 4-coordinated by oxygen. A reinvestigation of the chemistry of alkali-technetium-oxides formed under oxidizing conditions and at temperatures used to prepare nuclear waste glasses showed that higher coordinated alkali Tc(VII) oxide species had been reported, including those with the TcO5-and TcO6-anions. The chemistry of alkali Tc(VII) and other alkali-Tc-oxides is reviewed, along with relevant synthesis conditions.Additionally, we report attempts to make 5- and 6-coordinate pertechnetate compounds of K, Na, and Li, i.e. TcO5-and TcO6-. It was found that higher coordinated species are very sensitive to water, and easily decompose into their respective pertechnetates. It was difficult to obtain pure compounds, but mixtures of the pertechnetate and other phase(s) were frequently found, as evidenced by x-ray absorption spectroscopy (XAS), neutron diffraction (ND), and Raman spectroscopy. Low temperature electron paramagnetic resonance (EPR) measurements showed the possibility of Tc(IV) and Tc(VI) in Na3TcO5and Na5TcO6compounds.It was hypothesized that the smaller counter cation would result in more stable pertechnetates. To confirm the synthesis method, LiReO4and Li5ReO6were prepared, and their Raman spectra match those in the literature. Subsequently, the Tc versions LiTcO4and Li5TcO6were synthesized and characterized by ND, Raman spectroscopy, XANES, and EXAFS. The Li5TcO6was a marginally stable compound that appears to have the same structure as that known for Li5ReO6. Implications of the experimental work on stability of alkali technetate compounds and possible role in the volatilization of Tc are discussed.

The lead acceptor in p-type natural 2H-polytype MoS2 crystals evidenced by electron paramagnetic resonance

A low-temperature (T = 1.5–8 K) electron paramagnetic resonance study of p-type 2H-polytype natural MoS2 crystals reveals a previously unreported anisotropic signal of corresponding defect density (spin S = ½) ~5 × 1014 cm−3. For the applied magnetic field B//c-axis, the response is comprised of a single central asymmetric Zeeman peak at zero-crossing g = 2.102(1), amid a symmetrically positioned hyperfine doublet of splitting 6.6(2) G. Field angular observations reveal a two-branch g pattern, indicative of a defect of lower than axial symmetry, likely orthorhombic (C2v). Based on the signal specifics, it is ascribed to a system of decoupled Pb impurities substituting for Mo, the defect operating as an acceptor, with estimated thermal activation energy >10 meV. Supporting theoretical anticipation, the results pinpoint the conduct of the Pb impurity in layered MoS2.

Mechanically induced defects in milled CdZnSe nanocrystal: A low-temperature electron spin resonance spectroscopy analysis

The electron spin resonance (ESR) linewidth of the milled powder decreases as a function of increasing temperature. On the other hand, the intensity of the signal increases with increasing temperature from 108K to 243K and subsequently approaches Curie-like temperature behavior above 245K. These behaviors were understood in terms of the orbital degeneracy of fast-relaxing impurities, such as Fe3+ ion debris, from the grinding media that was embedded in the atomic structure of the milled powder and are not completely quenched. At lower temperature, only ±1/2 is occupied and the excited energy state is less likely effective in dipolar broadening compared with lower energy states of ±5/2 and ±3/2. In this regard, decrease in linewidth should be expected with increasing temperature.

Characterization and Subsequent Reactivity of an Fe-Peroxo Porphyrin Generated by Electrochemical Reductive Activation of O2.

Reductive activation of O2 is achieved by using the [FeIII(F20TPP)Cl] (F20TPP = 5,10,15,20-tetrakis(pentafluorophenyl) porphyrinate) porphyrin through electrochemical reduction of the [FeIII(F20TPP)(O2•-)] superoxo complex. Formation of the [FeIII(F20TPP)(OO)]- peroxo species is monitored by using low-temperature electronic absorption spectroscopy, electron paramagnetic resonance, and cyclic voltammetry. Its subsequent protonation to yield the [FeIII(F20TPP)(OOH)] hydroperoxo intermediate is probed using low-temperature electronic absorption spectroscopy and electron paramagnetic resonance.

Effect of Rabi splitting on the low-temperature electron paramagnetic resonance signal of anthracite

Specific distortions of the EPR signal of bulk anthracite are observed at low temperatures. They are accompanied by variations in the microwave oscillator frequency and are explained by the manifestation of the Rabi splitting due to the strong coupling between electron spins and the cavity, combined with the use of an automatic frequency-control (AFC) system. EPR signals are recorded at negligible saturation in the temperature range of 4–300K with use of the AFC system to keep the oscillator frequency locked to the resonant frequency of the TM110 cylinder cavity loaded with the sample. For the sample with a mass of 3.6mg the line distortions are observed below 50K and increase with temperature lowering. The oscillator frequency variations are used to estimate the coupling strength as well as the number of spins in the sample. It is shown that the spin-cavity coupling strength is inversely proportional to temperature and can be used for the absolute determination of the number of spins in a sample. Our results indicate that at low temperatures even 1016 spins of the anthracite sample, with a mass of about 0.5mg, can distort the EPR line.

261 - Mechanism of Protection by the Heavy Metal Poisoning Antidote 2,3-Dimercaptosuccinic Acid Against Copper-Induced DNA Cleavage Via Forming A Redox-Inactive Copper Complex

2,3-Dimercaptosuccinic acid (DMSA) has been widely used in the treatment of lead, arsenic and other heavy metal poisoning in clinic because of its low toxicity, safe and convenient administration. However, it is not clear whether DMSA can provide protection against copper-induced oxidative damage; and if so, what is its underlying molecular mechanism. In this study, we investigated the role of DMSA in copper-induced oxidative damage to DNA, using copper-containing system Cu(II)/ascorbate. Oxidative damage to DNA was measured as stand breakage and the formation of 8-oxodG. DMSA (0.1-1.0 mM) provided strong and dose-dependent protection against oxidation of DNA by Cu(II)/ascorbate system. In contrast, only limited protection was observed with the typical ●OH scavenger dimethyl sulfoxide, even at concentrations as high as 500 mM. DMSA also significantly inhibited copper-catalyzed oxidation of ascorbate and competed effectively with histidine and 1,10-phenanthroline for binding of cuprous copper, but not cupric copper, as demonstrated by UV-visible and low-temperature electron spin resonance techniques. We conclude that the protection by DMSA against copper-mediated oxidative damage to DNA is through the formation of a redox-inactive DMSA-cuprous complex.

ESR study of p-type natural 2H-polytype MoS2 crystals: The As acceptor activity

Low-temperature (T = 1.7–77 K) multi frequency electron spin resonance (ESR) study on p-type 2H-polytype geological MoS2 crystals reveals p-type doping predominantly originating from As atoms substituting for S sites in densities of (2.4 ± 0.2) × 1017 cm−3. Observation of a “half field”(g ∼ 3.88) signal firmly correlating with the central Zeeman As accepter signal indicates the presence of spin S &amp;gt; ½ As agglomerates, which together with the distinct multicomponent makeup of the Zeeman signal points to manifest non-uniform As doping; only ∼13% of the total As response originates from individual decoupled As dopants. From ESR monitoring the latter vs. T, an activation energy Ea = (0.7 ± 0.2) meV is obtained. This unveils As as a noticeable shallow acceptor dopant, appropriate for realization of effective p-type doping in targeted 2D MoS2-based switching devices.

Incorporation of pervasive impurities on HVPE GaN growth directions

High crystallinity thick films with low free carrier concentration (≤1×1015/cm3) and low compensation are required for many GaN-based electronic device applications. It has been demonstrated that low pressure chemical vapor and molecular beam epitaxy techniques can reproducibility deposit homoepitaxial films with low residual impurity concentrations. However, their typical slow growth rates prevent their utilization for thick film growth. Presently, hydride vapor phase epitaxy is the sole method that can deposit films with residual impurity concentrations ≤5×1016/cm3 at hundreds of microns per hour growth rate. It is crucial to verify if this method can reproducibly deliver thick free-standing GaN films of high crystalline quality with exceptionally low and uniform free carrier concentration. X-ray diffraction, Raman scattering, and low temperature photoluminescence experiments were carried out on a number of samples prepared by dicing a free-standing wafer into several pieces perpendicular and parallel to the major growth directions; namely, c-plane {0001}, a-plane {11−20}, and m-plane {1−100}. SIMS depth profiles were employed to identify and quantify the concentration of the pervasive impurities. Spatial maps of a Raman line sensitive to free-carrier concentration were measured to determine the spatial distribution of the net impurity concentration. The reduced concentration of un-compensated shallow donors was also verified by low temperature electron paramagnetic resonance.