Bruker Instruments Research Articles

A 3D-Printed Offline Nano-ESI Source for Bruker MS Instruments.

Nanoelectrospray ionization (nano-ESI) is a highly efficient and a widely used technique for the ionization of minute amounts of analyte. Offline nano-ESI sources are convenient for the direct infusion of complex mixtures that suffer from high matrix content and are crucial for the native mass spectrometric analysis of proteins. For Bruker instruments, no such source is readily available. Here we close this gap and present a 3D-printable nano-ESI source for Bruker instruments, which can be assembled by anyone with access to 3D printers. The source can be fitted to any Bruker mass spectrometer with an ionBooster ESI source and only requires minor, reversible changes to the original Bruker hardware. The general utility was demonstrated by recording high-resolution MS spectra of small molecules, intact proteins, as well as complex biological samples in negative and positive ion mode on two different Bruker instruments.

Python-based dynamic scheduling assistant for atmospheric measurements by Bruker instruments using OPUS.

Atmospheric remote sensing by instruments such as spectrometers and interferometers often requires scheduling that is dependent on external factors, for example; time and solar (or lunar) zenith angle. Such instruments manufactured by Bruker often use the software package OPUS, which, while useful, is not flexible enough for automatic, repeated, atmospheric measurements of this nature. In this brief paper, we describe ASAP, a Python tool developed to run our network of Fourier transform interferometers in New Zealand and Antarctica. It allows the automated scheduling of measurements by time, lunar, or solar zenith angle while accounting for weather or other external parameters. There is a wide range of useful functions, all packaged in a simple graphical user interface; it is available on request.

Deuterium nuclear magnetic resonance unambiguously quantifies pore and collagen-bound water in cortical bone

Bone water (BW) plays a pivotal role in nutrient transport and conferring bone with its viscoelastic mechanical properties. BW is partitioned between the pore spaces of the Haversian and lacuno-canalicular system, and water predominantly bound to the matrix proteins (essentially collagen). The general model of BW is that the former predominantly experiences fast isotropic molecular reorientation, whereas water in the bone matrix undergoes slower anisotropic rotational diffusion. Here, we provide direct evidence for the correctness of this model and show that unambiguous quantification in situ of these two functionally and dynamically different BW fractions is possible. The approach chosen relies on nuclear magnetic resonance (NMR) of deuterium ((2) H) that unambiguously separates and quantifies the two fractions on the basis of their distinguishing microdynamic properties. Twenty-four specimens of the human tibial cortex from 6 donors (3 male, 3 female, ages 27-83 years) were cored and (2) H spectra recorded at 62 MHz (9.4 Tesla) on a Bruker Instruments DMX 400 spectrometer after exchange of native BW with (2) H(2) O. Spectra consisted of a doublet signal resulting from quadrupole interaction of water bound to collagen. Doublet splittings were found to depend on the orientation of the osteonal axis with respect to the magnetic field direction (8.2 and 4.3 kHz for parallel and perpendicular orientation, respectively). In contrast, the isotropically reorienting pore-resident water yielded a single resonance line superimposed on the doublet. Nulling of the singlet resonance allowed separation of the two fractions. The results indicate that in human cortical bone 60% to 80% of detectable BW is collagen-bound. Porosity determined as the difference between total BW and collagen bound water fraction was found to strongly parallel micro-computed tomography (µCT)-based measurements (R(2) = 0.91). Our method provides means for direct validation of emerging relaxation-based measurements of cortical bone porosity by proton MRI.

Nearline acquisition and processing of liquid chromatography-tandem mass spectrometry data

Liquid chromatography–mass spectrometry (LC–MS) is a commonly used analytical platform for non-targeted metabolite profiling experiments. Although data acquisition, processing and statistical analyses are almost routine in such experiments, further annotation and subsequent identification of chemical compounds are not. For identification, tandem mass spectra provide valuable information towards the structure of chemical compounds. These are typically acquired online, in data-dependent mode, or offline, using handcrafted acquisition methods and manually extracted from raw data. Here, we present several methods to fast-track and improve both the acquisition and processing of LC–MS/MS data. Our nearly online (nearline) data-dependent tandem MS strategy creates a minimal set of LC–MS/MS acquisition methods for relevant features revealed by a preceding non-targeted profiling experiment. Using different filtering criteria, such as intensity or ion type, the acquisition of irrelevant spectra is minimized. Afterwards, LC–MS/MS raw data are processed with feature detection and grouping algorithms. The extracted tandem mass spectra can be used for both library search and de-novo identification methods. The algorithms are implemented in the R package MetShot and support the export to Bruker, Agilent or Waters QTOF instruments and the vendor-independent TraML standard. We evaluate the performance of our workflow on a Bruker micrOTOF-Q by comparison of automatically acquired and extracted tandem mass spectra obtained from a mixture of natural product standards against manually extracted reference spectra. Using Arabidopsis thaliana wild-type and biosynthetic gene knockout plants, we characterize the metabolic products of a biosynthetic pathway and demonstrate the integration of our approach into a typical non-targeted metabolite profiling workflow.

Phosphorylation at Ser-129 but Not the Phosphomimics S129E/D Inhibits the Fibrillation of α-Synuclein

alpha-Synuclein (alpha-syn) phosphorylation at serine 129 is characteristic of Parkinson disease (PD) and related alpha-synulceinopathies. However, whether phosphorylation promotes or inhibits alpha-syn aggregation and neurotoxicity in vivo remains unknown. This understanding is critical for elucidating the role of alpha-syn in the pathogenesis of PD and for development of therapeutic strategies for PD. To better understand the structural and molecular consequences of Ser-129 phosphorylation, we compared the biochemical, structural, and membrane binding properties of wild type alpha-syn to those of the phosphorylation mimics (S129E, S129D) as well as of in vitro phosphorylated alpha-syn using a battery of biophysical techniques. Our results demonstrate that phosphorylation at Ser-129 increases the conformational flexibility of alpha-syn and inhibits its fibrillogenesis in vitro but does not perturb its membrane-bound conformation. In addition, we show that the phosphorylation mimics (S129E/D) do not reproduce the effect of phosphorylation on the structural and aggregation properties of alpha-syn in vitro. Our findings have significant implications for current strategies to elucidate the role of phosphorylation in modulating protein structure and function in health and disease and provide novel insight into the underlying mechanisms that govern alpha-syn aggregation and toxicity in PD and related alpha-synulceinopathies.

Heterotrimeric G-protein α-Subunit Adopts a “Preactivated” Conformation When Associated with βγ-Subunits

Heterotrimeric G-protein α-Subunit Adopts a “Preactivated” Conformation When Associated with βγ-Subunits

Reconstitution and Characterization of Aminopyrrolnitrin Oxygenase, a Rieske N-Oxygenase That Catalyzes Unusual Arylamine Oxidation

Rieske oxygenases catalyze a wide variety of important oxidation reactions. Here we report the characterization of a novel Rieske N-oxygenase, aminopyrrolnitrin oxygenase (PrnD) that catalyzes the unusual oxidation of an arylamine to an arylnitro group. PrnD from Pseudomonas fluorescens Pf5 was functionally expressed in Escherichia coli, and the activity of the purified PrnD was reconstituted, which required in vitro assembly of the Rieske iron-sulfur cluster into the protein and the presence of NADPH, FMN, and an E. coli flavin reductase SsuE. Biochemical and bioinformatics studies indicated that the reconstituted PrnD contains a Rieske iron-sulfur cluster and a mononuclear iron center that are formed by residues Cys(69), Cys(88), His(71), His(91), Asp(323), His(186), and His(191), respectively. The enzyme showed a limited range of substrate specificity and catalyzed the conversion of aminopyrrolnitrin into pyrrolnitrin with K(m) = 191 microM and k(cat) = 6.8 min(-1). Isotope labeling experiments with (18)O(2) and H(2)(18)O suggested that the oxygen atoms in the pyrrolnitrin product are derived exclusively from molecular oxygen. In addition, it was found that the oxygenation of the arylamine substrates catalyzed by PrnD occurs at the enzyme active site and does not involve free radical chain reactions. By analogy to known examples of arylamine oxidation, a catalytic mechanism for the bioconversion of amino pyrrolnitrin into pyrrolnitrin was proposed. Our results should facilitate further mechanistic and crystallographic studies of this arylamine oxygenase and may provide a new enzymatic route for the synthesis of aromatic nitro compounds from their corresponding aromatic amines.

A Five-coordinate Metal Center in Co(II)-substituted VanX

In an effort to structurally probe the metal binding site in VanX, electronic absorption, EPR, and extended x-ray absorption fine structure (EXAFS) spectroscopic studies were conducted on Co(II)-substituted VanX. Electronic spectroscopy revealed the presence of Co(II) ligand field transitions that had molar absorptivities of approximately 100 m(-1) cm(-1), which suggests that Co(II) is five-coordinate in Co(II)-substituted VanX. Low temperature EPR spectra of Co(II)-substituted VanX were simulated using spin Hamiltonian parameters of M(S) = |+/-1/2), E/D = 0.14, g(real(x,y)) = 2.37, and g(real(z)) = 2.03. These parameters lead to the prediction that Co(II) in the enzyme is five-coordinate and that there may be at least one solvent-derived ligand. Single scattering fits of EXAFS data indicate that the metal ions in both native Zn(II)-containing and Co(II)-substituted VanX have the same coordination number and that the metal ions are coordinated by 5 nitrogen/oxygen ligands at approximately 2.0 angstroms. These data demonstrate that Co(II) (and Zn(II) from EXAFS studies) is five-coordinate in VanX in contrast to previous crystallographic studies (Bussiere, D. E., Pratt, S. D., Katz, L., Severin, J. M., Holzman, T., and Park, C. H. (1998) Mol. Cell 2, 75-84). These spectroscopic studies also demonstrate that the metal ion in Co(II)-substituted VanX when complexed with a phosphinate analog of substrate D-Ala-D-Ala is also five-coordinate.

Felix W. Wehrli, Editor‐in‐Chief: 13 years of dedicated service to Magnetic Resonance in Medicine

Felix W. Wehrli, Editor‐in‐Chief: 13 years of dedicated service to <i>Magnetic Resonance in Medicine</i>

NMR analysis of mixtures using hyphenation techniques and software

Two approaches to NMR analysis of mixtures used in Bruker instruments are considered: (1) a combination of high-performance liquid chromatography, NMR, and mass spectrometry, (2) the use of specialized software and spectral databases.

Application of Flow NMR to an Open-Access Pharmaceutical Environment

We present the application of flow NMR in an automated, open-access environment. The adjustment of parameters affecting the selection of the correct sample size, the elimination of carry-over, and the optimization of sample recovery are addressed. Advantages of this method include ease of use, elimination of NMR tubes, an delimination of handling errors that can result in the contamination of the probe. Sample throughput is similar to instruments using a conventional autosampler (for Bruker instruments, a BACS) although the time used for shimming the sample can be eliminated when the sample solvent does not change. The key feature of our methodology is that only one push solvent is used. This has major advantages over methods that switch solvents because there is no extensive flushing required between solvents and the deuterated push solvent, deuterium oxide, is economical. The disadvantage is the need to maintain the push and transfer solvents and usually the loss of the exchangeable protons that the sample may have. The protocol we present, using a single push solvent, contributes to the application of flow NMR in hands-on medicinal chemistry environments.

Phased array imaging on a 4.7T/33cm animal research system

Although phased array technology has been standard on clinical magnetic resonance imaging (MRI) systems for many years, it is just now becoming available on small animal MRI systems. Bruker Instruments Ltd. has recently provided multiple rf channels on a 4.7T/33cm Avance, but further development was necessary to complete the phased array implementation. This work discusses the development of this other hardware, specifically the rf array coil, low impedance preamplifiers, excitation coil, and image combination algorithm. Comparison of the array coil to a quadrature coil indicates superior signal-to-noise ratio from the array. In vivo images of a cat spine acquired simultaneously from the individual channels of the array and a sum of squares combination are shown.

Optimized Excitation and Automation for High-Resolution NMR Using B1-Insensitive Rotation Pulses

ADVERTISEMENT RETURN TO ISSUEPREVCommunicationNEXTOptimized Excitation and Automation for High-Resolution NMR Using B1-Insensitive Rotation PulsesPeter C. M. van Zijl, Tsang-Lin Hwang, Mark O'Neil Johnson, and Michael GarwoodView Author Information Departments of Radiology and Biophysics and Biophysical Chemistry Johns Hopkins University School of Medicine 217 Traylor Building, 720 Rutland Avenue Baltimore, Maryland 21205 Bruker Instruments, Inc., 47697 Westinghouse Drive Fremont, California 94539 Center for Magnetic Resonance Research University of Minnesota Minneapolis, Minnesota 55455 Cite this: J. Am. Chem. Soc. 1996, 118, 23, 5510–5511Publication Date (Web):June 12, 1996Publication History Received25 January 1996Published online12 June 1996Published inissue 1 January 1996https://doi.org/10.1021/ja9602612Copyright © 1996 American Chemical SocietyRIGHTS & PERMISSIONSArticle Views278Altmetric-Citations23LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit Read OnlinePDF (95 KB) Get e-AlertsSUBJECTS:Deposition,Optimization,Power Get e-Alerts

Spiral imaging on a small-bore system at 4.7T.

Spiral imaging has a number of advantages for ultrafast data acquisition. However, implementation on high-field small-bore systems requires carefully addressing the issues of inhomogeneity-induced blurring and gradient hardware constraints. In this paper, spiral imaging on a 40-cm-bore 4.7T CSI Omega System (Bruker Instruments) is discussed. A constant-voltage gradient waveform design algorithm is developed to reduce readout times as well as minimize waveform distortions due to gradient amplifier nonlinearities. Residual errors are then measured and taken into account in the image reconstruction procedure. Multiple spiral interleaves as well as a multifrequency reconstruction algorithm are used to decrease blurring of off-resonance spins. Both phantom and in vivo images demonstrate the performance of the resulting pulse sequences.

6] Modifications of older model nuclear magnetic resonance console for collection of multinuclear, multidimensional spectral data

Pursuant to the original description by Kay, Ikura, Tschudin, and Bax, a growing variety of double- and triple-resonance, three-dimensional (3D) and four-dimensional (4D) nuclear magnetic resonance (NMR) experiments have been developed. Although fully equipped commercial NMR spectrometer consoles produced after 1990 have been capable of performing these demanding experiments, earlier consoles lack the necessary decoupling capabilities and requisite number of transmitter channels. Thus, although the original two-dimensional (2D) triple-resonance experiments have been able to be carried out some of with a standard 1985 vintage Bruker Instruments AM-400 NMR spectrometer and BSV-3 X-nucleus decoupler, it was needed to make extensive modifications in order to perform multinuclear 3D and 4D experiments. Kay et al. described the adaptation of a Bruker AM console for early 3D and 4D triple-resonance experiments. With more extensive modifications, most of the current multinuclear 3D and 4D experiments have been able to be performed with older model Bruker Instruments AM-500 and AM-600 consoles and BSV-3 X-nucleus decouplers.

Short Communication: Intracellular pH Decreases During Entry into Estivation in the Land Snail Oreohelix Strigosa

Given that respiratory acidosis occurs in land snails during entry into estivation (for a review, see Barnhart, 1989), it is of interest to know the extent of intracellular acidification during this transition. Efforts to measure intracellular pH (pHi) of estivating snails, however, have been complicated by the fact that manipulations inherent in some techniques (e.g. injection of radiolabeled tracers) may cause arousal of dormant snails (B. B. Rees, personal observations). In the present study, we report on the use of P-n.m.r. as a non-invasive technique for measuring pHi in the land snail Oreohelix strigosa (Gould) during entry into estivation. This approach has allowed us to obtain a continuous record of pHi during this transition without disturbance to the animal. We have found that pHi declines rapidly and that over 3 days of estivation pHi drops by 0.4-0.5 units. Since the metabolism of a variety of organisms and cell types is sensitive to pHi (Busa and Nuccitelli, 1984), alterations of pHi during estivation may contribute to the overall metabolic suppression observed in this quiescent state. O. strigosa were collected near Glenwood Springs, Colorado, in November 1990 and were maintained in the laboratory as previously described (Rees and Hand, 1990). The whole mass (soft tissue plus shell) of the snails ranged from 1.0 to 1.5 g. Following the n.m.r. experiments, species identification was confirmed electrophoretically (Rees, 1988). n.m.r. studies were performed on a 7.05-T 10-cm bore cryomagnet with an AM300 spectrometer (Bruker Instruments, Billerica, MA) using a home-built two-turn surface coil. The surface coil was made of insulated 2 mm copper wire and had a diameter of lcm. Fixed and variable capacitors (RS Laboratories, Oxford, UK) were placed for tuning and balanced matching. This coil had a loaded Q that exceeded 100. Approximately 30min prior to the start of an experiment, estivating snails were aroused by transferring them to dampened paper toweling. Snails were then

Nuclear magnetic resonance evaluation of stroke. A preliminary report.

Nine patients who had acute and subacute stroke were examined by nuclear magnetic resonance (NMR) using a 6-MHz Bruker Instruments proton scanner. A modified Carr-Purcell-Meiboom-Gill pulse sequence was used for signal detection. The resultant string of spin-echoes was Fourier transformed into projections that were subsequently back-projected to a series of spin-echo images. From these images, spin density and T2 were calculated for each pixel. The NMR scans revealed stroke in each of the patients, while CT demonstrated only eight of the lesions. T2 was prolonged in all of the ischemic regions and is the most sensitive NMR parameter in detecting stroke. These preliminary results suggest that NMR scanning of patients who have acute stroke may be clinically useful, and that the T2 component of the NMR signal is most important.

Improvement in the cross-polarization NMR experiment for suppression of rigid protonated carbons

.4 very useful NMR experiment for solids simply requires the use of a calculated delay for the decoupling pulse (1). For a proper choice of the decoupler delay TD, singly or doubly protonated carbons can usually be suppressed from the carbon13 NMR spectrum. Owing to their usually rapid motion at room temperatures, triply protonated carbons (methyl groups) are usually not suppressed as easily. This experiment has received various titles such as “dipolar dephasing” or “interrupted decoupler.” It has been used by various researchers to simplify complicated carbon13 spectra of biological molecules (2, 3), to estimate carbon types in fuels (4), and to substantiate chemical reaction mechanisms (5). One problem with this experiment is that the decoupler delay time TD introduces an additional linear phase error in the observed NMR spectrum if the signal digitizer is triggered at the end of the delay. Alternately, if the digitization is begun prior to the delay, the additional linear phase error can be avoided at the expense of possibly recording (1) the would-be suppressed carbon signals and (2) an rf transient when the decoupler pulse is activated. If a chemical-shift refocusing pulse is added to the rare-spin pulse sequence at the center of the decoupler delay TD, then the digitization can begin at the end of the delay without the undesired linear phase error. The pulse sequence is depicted in Fig. 1. Also shown are the appropriate rf rotating-frame phases for a 4-cycle phase alternated quadrature signal averaging. The rare-spin FID is inverted by alternating the spin-lock direction of the abundant spin (X, x), (6, 7). The quadrature outputs of the real (R) and imaginary (I) channels of the digitizer are added/subtracted from the even (E) and odd (0) memory locations as described in Fig. 1. Carbon13 measurements were made on a commercially available (Aldrich) powder of pdi-tert-butylbenzene (PDTBB). The proton T, of this compound was measured as approximately 0.3 sec. Solids spectra were recorded with an IBM Instruments, Inc. 100 MHz Solids Accessory and a Bruker Instruments, Inc. WPlOOSY console. The carbon-l 3 spectra shown in Fig. 2 were recorded at 25.18 MHz using approximately 300 mg of the powder, a 4.8 kHz MAS rotor speed, a cross-polarization contact time of 2 msec, a spectral width of 15 kHz, a filter width of 19 kHz, a repetition time of 3 set, and 48 signal averages. The matched Hartmann-Hahn and decoupling rf fields were 42 kHz. The width of the ?r refocusing pulse on the carbon13 channel was 12