Reaction Interface Mass Spectrometry Research Articles

Nitrogen Trifluoride: A New Reactant Gas in Chemical Reaction Interface Mass Spectrometry for Detection of Phosphorus, Deuterium, Chlorine, and Sulfur

Nitrogen Trifluoride: A New Reactant Gas in Chemical Reaction Interface Mass Spectrometry for Detection of Phosphorus, Deuterium, Chlorine, and Sulfur

Chemical Reaction Interface Mass Spectrometry with High Efficiency Nebulization

A high efficiency nebulizer (HEN) coupled to a heated spray chamber and a membrane desolvator is used for liquid sample introduction in chemical reaction interface mass spectrometry (CRIMS). Compared to the conventional thermospray nebulizer operated at solvent flow rate of 1 mL/min, the HEN provides small droplets at lower flow rates (10-100 microL/min), improving the desolvation and analyte transport efficiency. As a result, the sensitivity for carbon detection by CRIMS is improved by a factor of 4. The new arrangement offers an easy-to-use and robust interface, facilitating the availability of a variety of liquid chromatographic techniques to the CRIMS. Separation and detection of labeled peptides in a mixture of unlabeled biopolymers is illustrated at a solvent flow rate of 45 microL/min as an example of new possibilities offered by the improved liquid introduction interface.

Detection of sulfur-containing impurities in pharmaceutical samples by high performance liquid chromatography/chemical reaction interface mass spectrometry.

This paper describes the use of chemical reaction interface mass spectrometry (CRIMS) combined with liquid chromatography for the detection of trace level sulfur-containing impurities in pharmaceutical materials. A mixture of sulfur- and non-sulfur-containing compounds were analyzed initially to test the system. Then the determination of trace level impurities in a cimetidine drug substance was carried out. Detection of sulfur-containing impurities at less than 0.1% of the major component was obtained with good linearity. The results obtained are consistent with the expected results for this sample and illustrate the applicability of the technique.

Mass spectrometry innovations in drug discovery and development.

This review highlights the many roles mass spectrometry plays in the discovery and development of new therapeutics by both the pharmaceutical and the biotechnology industries. Innovations in mass spectrometer source design, improvements to mass accuracy, and implementation of computer-controlled automation have accelerated the purification and characterization of compounds derived from combinatorial libraries, as well as the throughput of pharmacokinetics studies. The use of accelerator mass spectrometry, chemical reaction interface-mass spectrometry and continuous flow-isotope ratio mass spectrometry are promising alternatives for conducting mass balance studies in man. To meet the technical challenges of proteomics, discovery groups in biotechnology companies have led the way to development of instruments with greater sensitivity and mass accuracy (e.g., MALDI-TOF, ESI-Q-TOF, Ion Trap), the miniaturization of separation techniques and ion sources (e.g., capillary HPLC and nanospray), and the utilization of bioinformatics. Affinity-based methods coupled to mass spectrometry are allowing rapid and selective identification of both synthetic and biological molecules. With decreasing instrument cost and size and increasing reliability, mass spectrometers are penetrating both the manufacturing and the quality control arenas. The next generation of technologies to simplify the investigation of the complex fate of novel pharmaceutical entities in vitro and in vivo will be chip-based approaches coupled with mass spectrometry.

An innovative method for measuring hydrogen and deuterium: chemical reaction interface mass spectrometry with nitrogen reactant gas

A new method for measuring deuterium isotopic enrichment with CRIMS (chemical reaction interface mass spectrometry) is described. Using nitrogen as the reactant gas in a chemical reaction interface generates molecular hydrogen that provides the H 2 and HD from which the deuterium content can be analyzed with a benchtop quadrupole mass spectrometer. Samples of deuterated leucine in unlabeled leucine were used as the primary test species. Detection of deuterium enrichment was accurate, precise, and linear. We used this scheme to evaluate the results of a process to acetylate lysine residues in a peptide–neurotensin. With separation on a C 18 column, we found a 61% yield of the desired monoethylated product that had a D/H ratio very close to the theoretical one. Isotope ratio monitoring for deuterated species will be important in metabolism studies where CRIMS generates a comprehensive and quantitative view of products of deuterated precursors. Where concerns about metabolic isotope effects of deuterium are absent, the use of deuterium will enable these studies to be performed with simpler syntheses and at less cost than if using 13C or 15N.

Size exclusion chromatography-chemical reaction interface mass spectrometry: "a perfect match".

Size exclusion chromatography (SEC) has been coupled to chemical reaction interface mass spectrometry (CRIMS) for the analysis of biopolymers. This innovative combination allows the analysis of biopolymers with no limitation on the molecular weight and chemical composition of the species under investigation. With SEC-CRIMS we have examined different classes of biopolymers including polynucleotides, proteins, and polysaccharides. Moreover, CRIMS allows the simultaneous detection of multiple organic elements and their stable isotopes. When SEC is interfaced to CRIMS, further information (elemental detection) is obtained, enhancing the analytical capability of SEC. These features have been applied to the detection of a labeled protein (13C-rat growth hormone) in plasma and to the characterization of heparin and low molecular weight heparin from different sources.

Pharmacokinetic and metabolism studies using uniformly stable isotope labeled proteins with HPLC/CRIMS detection.

We present a novel method for performing pharmacokinetic and metabolism studies on macromolecules that offers advantages over the existing techniques of radiolabeling, immunoassay or bioassays. Our strategy uses macromolecules with stable isotopes uniformly distributed throughout the structure. The stable isotope enrichment is detected using high performance liquid chromatography combined with chemical reaction interface mass spectrometry (HPLC/CRIMS). HPLC/CRIMS is a technique where analytes are first eluted from an HPLC column and then dissociated in a microwave reaction chamber. The dissociated analytes are oxidized using SO2 and the resulting small molecules are detected by the mass spectrometer. The stable-isotope labeled analyte is distinguished from the matrix carbon by monitoring the enrichment of 13CO2. In order to demonstrate the feasibility of performing pharmacokinetic and metabolism studies using this technique, uniformly 13C, 15N-labeled rat growth hormone was administered intravenously to rats and blood samples were collected. Raw plasma samples were analysed by HPLC/CRIMS. Growth hormone was detectable for 1 h following administration. The absolute amounts detected ranged from a high of 66 pmol to a low of 825 fmol in a 20 microL plasma sample. The data were modeled using PCNONLIN and were consistent with a one compartment model. The calculated half-life was 7.7 +/- 0.7 min, with a clearance of 4.5 +/- 0.3 mL min(-1), values consistent with literature reports for growth hormone in rats. No circulating growth hormone metabolites were detected in the plasma. This paper demonstrates a novel technique for performing pharmacokinetic studies of proteins. The uniform labeling strategy also presents a viable comprehensive method for obtaining metabolism data on macromolecules.

Dissociative electron attachment negative ion mass spectrometry: a chlorine-specific detector for gas chromatography

This work describes the application of negative ion chemical ionization, optimized for dissociative electron attachment (DEA), to location of unknown trace chlorinated compounds in complex gas chromatograms by selected ion recording (SIR) of m / z 35 and 37. The DEA-SIR technique is compared with other GC detectors, including the electron capture detector, electrolytic conductivity detector, the atomic emission detector and the chemical reaction interface mass spectrometry method, with respect to selectivity for chlorine, sensitivity, linear dynamic range, and general robustness and ease of use. When applied to quantitative analysis of target analytes such as polychlorobiphenyls, the DEA-SIR method has potential problems arising from the possibility of suppression effects due to abundant co-eluting components, and possible alleviating measures are discussed. In addition to these practical investigations, literature information on the fundamental physical and chemical phenomena underlying the DEA process is summarized in order to guide future work on extension to other compound types and on general improvements to the technique.

Selective Detection of Selenium in Water Utilizing Chemical Reaction Interface Mass Spectrometry†

Gas chromatography/chemical reaction interface mass spectrometry (GC/CRIMS) is shown to be a successful selective method for the detection of selenium-containing compounds. Two reaction gases, sulfur dioxide (SO 2 ) and hydrogen chloride (HCl), were examined in order to optimize selectivity and sensitivity. A high degree of selectivity was obtained with SO 2 as a reaction gas; however, the detection limit of 50 SeO 3 + at m/z 128 (the most sensitive ion for the SO 2 -containing plasma) was only 3 ng μl -1 . HCl gas, which had been shown to be a good reaction gas for snufur-containing compounds, was also shown to be an excellent reaction gas for selenium-containing compounds. In the HCl-containing plasma, 80 SeCl + at m/z 115 was the most sensitive and selective ion for the detection of selenium-containing compounds. Selectivity was demonstrated by using mixtures of selenium-containing and non-selenium-containing compounds. The utility of GC/CRIMS as a method for the selective detection of selenium-containing compounds was demonstrated with a variety of selenium complexes that were formed by the addition of selective selenium complexing agents to selenium-containing water. The detection limit of selenium in water was ∼62 pg and the linear dynamic range spanned at least two orders of magnitude (620 pg pl -1 -308 ng μl -1 ).

ChemInform Abstract: Liquid Chromatography/Chemical Reaction Interface Mass Spectrometry as an Alternative to Radioisotopes for Quantitative Drug Metabolism Studies.

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Liquid chromatography/chemical reaction interface mass spectrometry as an alternative to radioisotopes for quantitative drug metabolism studies.

Chemical reaction interface mass spectrometry (CRIMS) was coupled on-line with HPLC using a Vestec particle beam interface. A helium-assisted nebulizer provided added stability with no loss in accuracy or precision as compared to the thermospray nebulizer at flow rates of up to 1.0 mL/min using isocratic conditions. However, mass spectral response was found to be solvent-dependent for both the helium-assisted and thermospray nebulizers. Postcolumn solvent addition of methanol eliminated solvent-dependent decreases in mass spectral response. This allowed gradient HPLC elutions to be performed. Under these conditions, the flow of solvent into the particle beam interface was 2.5 mL/min, so a conventional thermospray nebulizer had to be used instead of the helium-assisted nebulizer. Experiments were conducted with the antianxiety agent buspirone in order to validate the methodology. Metabolites from in vitro incubations of [15N]/[14C]buspirone with rat liver slices were analyzed by gradient LC/CRIMS and by gradient LC/[14C] radioactivity counting. The response from LC/CRIMS analysis for individual metabolites was then compared with that obtained by LC/[14C] radioactivity counting. An excellent correlation was observed between the two methods for metabolites with quite different HPLC characteristics. Thus, gradient LC/CRIMS in combination with stable isotopes provides an alternative to using radioisotopes for carrying out drug metabolism studies.

Element- and isotope-specific detection for high-performance liquid chromatography using chemical reaction interface mass spectrometry

We have developed a combination of high-performance liquid chromatography (HPLC) and the chemical reaction interface mass spectrometry (CRIMS) method by using a Vestec Universal Interface (UI). This interface provides the extremely high degree of solvent removal that the CRIMS process requires. In doing so, we have produced an HPLC detector with the ability to carry out element- and isotope-selective analyses with detection that is inherently: linear, structure-independent, sensitive, selective, comprehensive and flexible. The characteristics of the instrumentation and its performance are described.

Stable-Isotope Dilution Studies of an Intact Protein Using High-Performance Liquid Chromatography/Chemical Reaction Interface Mass Spectrometry

Stable-Isotope Dilution Studies of an Intact Protein Using High-Performance Liquid Chromatography/Chemical Reaction Interface Mass Spectrometry

Nitrogen Trifluoride: A New Reactant Gas in Chemical Reaction Interface Mass Spectrometry for Detection of Phosphorus, Deuterium, Chlorine, and Sulfur

A new set of reactions that involve fluorine have been investigated in chemical reaction interface mass spectrometry (CRIMS). The primary goal of this study was to provide a means to selectively detect phosphorus-containing compounds, which fluorine does by generating PF 5 . P F 4 + , the main fragment ion of PF 5 , provides a sensitive (10 pg/s), selective, and linear (> 10 5 dynamic range) channel for phosphorus-containing analytes. This fluorine-rich environment also provides new ways to selectively and simultaneously detect hydrogen isotopes, chlorine, and sulfur. NF 3 as a reactant gas provides the most comprehensive array of elemental and isotopic detection yet available for CRIMS. An application of phosphorus-selective detection was attempted by examination of a plasma sample from a patient who had been treated with cyclophosphamide. The phosphorus-selective channel showed six peaks: one is derivatized phosphate, two are t -butyldimethylsilyl (TBDMS) derivatives of cyclophosphamide, another two are TBDMS derivatives of 4-hydroxy-yclophosphamide, and one is from underivatized cyclophosphamide. The simultaneous detection of chlorinecontaining compounds confirmed our results for those peaks related to cyclophosphamide and its metabolite.

Direct determination of human urinary cortisol metabolites by [formula omitted

Feasibility of using high performance liquid chromatographic input to the chemical reaction interface mass spectrometry system was assessed by measuring the profile of hydrolyzed urinary metabolites of [9,12,12- 2H 3]cortisol in six human subjects with no preparation other than hydrolysis and solid phase extraction. Relative amounts of tetrahydrocortisol, tetrahydrocortisone, and cortolones (as the sum of α- and β-) were 0.417 ± 0.047, 0.523 ± 0.036 and 0.059 ± 0.019, respectively. The constant reproducibility of the measurements coupled with a profile consistent with that observed by other workers shows that the technique represents an important tool in the determination of metabolites of endogenous molecules.

Tracing 15N with chemical reaction interface mass spectrometry: a demonstration using 15N-labeled glutamine and asparagine substrates in cell culture.

This research demonstrates how the chemical reaction interface mass spectrometry (CRIMS) approach works for a study of amino acid metabolism in cell culture. 15N-selective chromatograms from both the culture medium and the cytosol of human hepatoma Hep G2 cells that were incubated in the presence of either 12 mM (alpha-15N)glutamine or (alpha-15N)asparagine have been produced. The time course of the distribution of 15N among different amino acids, as well as the enrichment for each amino acid, were observed over a 144 h period. Labeled glutamine was quickly converted into glutamate. After 144 h of incubation, the total amount of 15N was distributed primarily among alanine (50%), proline (28%) and glutamate (21%). The 15N enrichment of alanine and proline reached 44% and 41% respectively. Asparagine was only slowly metabolized by the cells. In addition to the 82% that was retained in asparagine, the remaining 15N in the media at 144 h was found primarily in alanine (8%), glutamate (6.8%) and proline (2.2%). Their enrichments were 20%, 36% and 19% respectively. The minimum detectable amount was 17 pg of 15N entering the CRI. CRIMS appears to be a powerful, facile approach for 15N-tracer experiments.

Application of high-performance liquid chromatography/chemical reaction interface mass spectrometry for the analysis of conjugated metabolites: a demonstration using deuterated acetaminophen.

The combination of a universal high-performance liquid chromatography/mass spectrometry (HPLC/MS) interface (UI) and the element and isotope-selective capabilities of the chemical reaction interface (CRI) has potential as a comprehensive analysis system for drug conjugates. In this work, we found equal sensitivity for model compounds as their sulfate or glucuronide conjugates. We examined urine and bile samples from Syrian golden hamsters after dosing with (2H4)acetaminophen (D4-APAP), with particular emphasis on the rich range of conjugated metabolites that are known to be produced. Seventeen metabolites were quantified from a single chromatogram of urine; 14 were conjugates. With a combination of authentic standards, selective hydrolysis, and sulfur-selective CRIMS detection, at least partial identification of most of these metabolites was accomplished. The glutathione conjugate of APAP appears the dominant metabolite in bile. The quantitative pattern of APAP metabolism found here is consistent with literature values. It does appear that this HPLC/UI/CRIMS combination has substantial ability to carry out comprehensive metabolite determinations, especially for conjugated species.

Implementation of the chemical reaction interface mass spectrometry technique on a Hewlett-Packard mass-selective detector

A microwave-powered chemical reaction interface has been installed in a Hewlett-Packard gas chromatograph-mass spectrometer (GC-MS) system (5890 II gas chromatograph-5971 mass-selective detector). The technical details and optimization strategies are discussed. The evaluation of this new setup is presented, showing detection limits of 1 ng of 13C-, 15N-, and Cl-containing compounds with signal-to-noise ratios greater than or equal to 3. Selective detection was evaluated with a urine sample from a dog dosed with 15N 3-midazolam that had been previously analyzed by using a differentially pumped research-level quadrupole mass spectrometer. The results show that the detection of 15N and Cl remains highly selective and the mass-selective detector gives comparable sensitivity to the larger instrument when the latter is operating over a conventional mass range. The capability for chemical reaction interface mass spectrometry can be easily accomplished with an inexpensive GC-MS system.

Selective detection of chlorine-containing compounds by gas chromatography/chemical reaction interface mass spectrometry

Chemical reaction interface mass spectrometry (CRIMS) was studied as a gas chromatographic detection technique for chlorine-containing compounds. Both SO2 and HBr were tested as reactant gases. With SO2, a detection limit of 50 pg of diazepam and a linear range of 4 orders of magnitude were achieved, and the experimental data were reproducible. With HBr, the detection limit was 10 ng of diazepam and the linear dynamic range was only 2 orders of magnitude. The possible pharmacological application of CRIMS was studied using urine spiked with diazepam and several of its metabolites, and the results show CRIMS to be a simple but potentially powerful method in drug metabolism studies.

Selective detection and characterization of chlorine- and bromine-containing compounds in complex mixtures using microwave-induced plasma/chemical reaction interface mass spectrometry

In the environmental and pharmacological sciences, it is important to selectively detect chlorine- and bromine-containing compounds in complex mixtures. Currently, a new technique called microwave-induced plasma/chemical reaction interface mass spectrometry (MIP/CRIMS) is being used as a selective detector of elements and stable isotopes. This technique, which involves post-column reactions (a reaction interface), includes a low-pressure microwave-induced helium plasma (MIP) to which a reaction gas is added. Effluents of a chromatographic column that enter this reaction interface are converted into small stable neutrals. The mass spectra of these neutrals will identify and quantify the elements and isotopes of interest. Once the retention times of the peaks of interest are obtained, their full mass spectra can be acquired by repeating the experiment with the MIP off. This method combines the sensitivity of a halogen specific detector with the compound identification of mass spectrometry. In this study, SO2 has been found highly effective as a reaction gas for selective detection of chlorine- and bromine-containing compounds using GC/MIP/CRIMS. Detection limits of 10 pg and 1 ng, and dynamic range of at least four and two orders of magnitude, were achieved for chlorine- and bromine-containing compounds, respectively. The selective detection of chlorinated compounds in complex mixtures is demonstrated by using a mixture of non-chlorinated compounds and polychlorinated biphenyls, and by selective detection of triclopyr (a chlorinated herbicide) in extracts of leaves of garden bean (Phaseolus vulgaris var. Topcrop).