Chemical ionization mass spectrometry II. Diferentiation of primary, secondary, and tertiary amines

Abstract. A systematic study was performed to investigate the utility of atmospheric pressure chemical ionization hydrogen-deuterium exchange mass spectrometry (APCI HDX MS) to identify the structures of nitrogen-containing aromatic compounds. First, experiments were performed to determine the optimized experimental conditions, with dichloromethane and CH 3 OD found to be good cosolvents for APCI HDX. In addition, a positive correlation between the heated capillary temperature and the observed HDX signal was observed, and it was suggested that the HDX reaction occurred when molecules were contained in the solvent cluster. Second, 20 standard nitrogen-containing compounds were analyzed to investigate whether speciation could be determined based on the different types of ions produced from nitrogencontaining compounds with various functional groups. The number of exchanges occurring within the compounds correlated well with the number of active hydrogen atoms attached to nitrogen, and it was confirmed that APCI HDX MS could be used to determine speciation. The results obtained by APCI HDX MS were combined with the subsequent investigation of the double bond equivalence distribution and indicated that resins of shale oil extract contained mostly pyridine type nitrogen compounds. This study confirmed that APCI HDX MS can be added to previously reported chemical ionization, electrospray ionization, and atmospheric pressure photo ionization-based HDX methods, which can be used for structural elucidation by mass spectrometry.

Section: Analysis Of Standard Compounds Containing Nitrogensupporting

confidence: 87%

Optimization and Application of APCI Hydrogen–Deuterium Exchange Mass Spectrometry (HDX MS) for the Speciation of Nitrogen Compounds

Acter

Cho

Kim

et al. 2015

“…This fragmentation pathway may proceed via a labile intermediate carbamoyl acid intermediate 3, which easily eliminates carbon dioxide and is therefore often not observed. Because the unprotected amino alcohol 4 affords fragment 2 after ionization in the MS detector, mass spectrometry does not provide a means to differentiate between 1 and 4, i.e., the determination of the presence of the t-Boc group requires isolation and further spectroscopic analysis or the use of reference samples for a time-consuming comparison of HPLC retention times which is unsuitable for high-throughput screening.Structure elucidation by mass spectrometry has become increasingly successful through the introduction of chemical ionization with ND 3 , D 2 O, and CD 3 OD for exchange of hydrogen for deuterium of organic compounds in the gas phase [2]. This technique can be applied to H/D exchange MS analysis of various classes of compounds including alcohols, carboxylic acids, amines, amides, and thiols [3].…”

mentioning

confidence: 99%

“…Structure elucidation by mass spectrometry has become increasingly successful through the introduction of chemical ionization with ND 3 , D 2 O, and CD 3 OD for exchange of hydrogen for deuterium of organic compounds in the gas phase [2]. This technique can be applied to H/D exchange MS analysis of various classes of compounds including alcohols, carboxylic acids, amines, amides, and thiols [3].…”

mentioning

confidence: 99%

Elucidation of the presence and location of t-Boc protecting groups in amines and dipeptides using on-column H/D exchange HPLC/ESI/MS

Wolf

Villalobos

Cummings

et al. 2005

High performance liquid chromatography/mass spectrometry (HPLC/MS) has become a widely used technique for routine analysis of pharmaceutical compounds. The constant search for new drugs requires the development of time-efficient methods that can be employed in high-throughput screening of combinatorial libraries of a variety of compounds, including amines and peptides. Conventional HPLC/MS is a powerful technique that can easily be automated and is suitable for comprehensive screening purposes. However, the unequivocal determination of the presence and location of important carbamoyl protecting groups of amines is often elusive because of their inherent instability under MS conditions. In this study, the use of on-column H/D exchange HPLC/ESI/MS for structure elucidation of t-Boc protecting groups which can often not be detected by MS because of facile McLafferty rearrangement has been examined. We demonstrate that employing a deuterated mobile phase in HPLC/MS analysis provides a convenient tool for the determination of the absence or presence of t-Boc protecting groups in amines and peptides. . This fragmentation pathway may proceed via a labile intermediate carbamoyl acid intermediate 3, which easily eliminates carbon dioxide and is therefore often not observed. Because the unprotected amino alcohol 4 affords fragment 2 after ionization in the MS detector, mass spectrometry does not provide a means to differentiate between 1 and 4, i.e., the determination of the presence of the t-Boc group requires isolation and further spectroscopic analysis or the use of reference samples for a time-consuming comparison of HPLC retention times which is unsuitable for high-throughput screening.Structure elucidation by mass spectrometry has become increasingly successful through the introduction of chemical ionization with ND 3 , D 2 O, and CD 3 OD for exchange of hydrogen for deuterium of organic compounds in the gas phase [2]. This technique can be applied to H/D exchange MS analysis of various classes of compounds including alcohols, carboxylic acids, amines, amides, and thiols [3]. It has also been shown that hydrogens bonded to carbon atoms in aromatic compounds can be replaced by deuterium in the gas phase [4]. The coupling of HPLC and mass spectrometry combines a powerful separation technique with the sensitive and informative MS detection mode, which opens a new venue for H/D exchange MS analysis of complex mixtures containing polar compounds. To date, various MS techniques including chemical ionization detection mass spectrometry (CI/MS) [5], fast atom

“…NMR methods, when coupled with H/D exchange are the ideal choice for monitoring individual residues or each amide hydrogen; however, these methods are limited to highly purified proteins or metabolites that are soluble at high concentrations, thus eliminating the possibility of determining structural features of drugs and metabolites that are in the discovery stage of drug development and are only available in limited quantities. On the contrary, mass spectrometry methods have advantages from inherent sensitivity and selectivity.As early as 1971, Hunt et al [9] showed that under chemical ionization (CI) conditions, H/D exchange can be rapidly achieved by using either CH 3 OD or ND 3 as reagent gas. Later, Hunt and Sethi [10] showed that the proton affinity difference between the analyte and the reagent gas is a major factor in determining the degree of exchange in chemical ionization mass spectrometry.…”

mentioning

confidence: 99%

“…As early as 1971, Hunt et al [9] showed that under chemical ionization (CI) conditions, H/D exchange can be rapidly achieved by using either CH 3 OD or ND 3 as reagent gas. Later, Hunt and Sethi [10] showed that the proton affinity difference between the analyte and the reagent gas is a major factor in determining the degree of exchange in chemical ionization mass spectrometry.…”

mentioning

confidence: 99%

In electrospray ionization source hydrogen/deuterium exchange LC-MS and LC-MS/MS for characterization of metabolites

Lam

Ramanathan

2002

122

A new method is described for performing hydrogen/deuterium (H/D) exchange in an electrospray ionization (ESI) source. The use of liquid chromatography (LC)-mass spectrometer equipped with an ESI source and deuterium oxide (D 2 O) as the sheath liquid allows H/D exchange experiments to be performed on-line. This directly provides information for determining the number and position of exchangeable hydrogens, aiding in the elucidation of the structures of drug metabolites. To demonstrate the utility of this method, LC-mass spectrometry (MS) and LC-MS/MS experiments were performed using either H 2 O or D 2 O as sheath liquid on a matrix metalloprotease (MMP) inhibitor (PD 0200126) and its metabolites. Examination of the mass shift of the deuteriated molecule from that of the protonated molecule allowed the number of exchangeable protons to be determined. Interpretation of the production-spectra helped to determine the location of the exchanged protons and assisted in the assignment of the site uring the process of drug discovery, it is highly desirable to increase the number of successful drug candidates for preclinical, clinical and commercial development. Therefore, the drug discovery process is constantly scrutinized and improved [1]. Adding to this pressure is the generation of vast numbers of new chemical entities resulting from combinatorial chemistry technology [2]. Drug metabolism plays an important role in the drug discovery process [3]. Specifically, the identification of metabolites during the early stage of development can be helpful to medicinal chemists trying to block some of the metabolic hot spots and produce an appropriate drug that is less susceptible to metabolism and increase the half-life of the drug. Therefore, rapid identification of drug metabolites is imperative for drug development [4,5].Hydrogen/deuterium exchange is a well-established technique for studying structure, stability, folding dynamics, and intermolecular interactions in proteins in solution [6]. During solution phase H/D exchange, labile protons in the side chains and amide hydrogens, which are not protected from solution generally exchange rapidly. Exchanges of these unprotected hydrogens occur on the order of a few to a few tenths per second under the experimental conditions described in the aforementioned studies. If however, amide or side chain hydrogens are protected from solution (e.g., when they are hydrogen-bonded in structurally stable secondary-structure elements), the exchange rates can be considerably reduced. Methods in which H/D exchange experiments are combined with either nuclear magnetic resonance (NMR) spectroscopy or mass spectrometry are also well-established [7,8]. NMR methods, when coupled with H/D exchange are the ideal choice for monitoring individual residues or each amide hydrogen; however, these methods are limited to highly purified proteins or metabolites that are soluble at high concentrations, thus eliminating the possibility of determining structural features of drugs and metabolites that are i...