A high performance low magnetic field internal electrospray ionization-fourier transform ion cyclotron resonance mass spectrometer

A new in-magnetic field electrospray ionization (ESI) and Fourier transform ion cyclotron resonance mass spectrometer has been constructed and evaluated. This system is characterized by the use of multiple concentric cryopanels to achieve ultrahigh vacuum in the ion cyclotron resonance cell region, a probe-mounted internal ESI source, and a novel in-field shutter. Initial experiments demonstrate high resolution mass measurement capability at a field strength of 1 T. Mass resolution of 700,000 has been obtained for the 3+ charge state of Met-Lys-bradykinin (at m/z 440) generated by electrospray ionization. When electron impact ionization was employed, resolution in excess of 9,200,000 was achieved for nitrogen molecular ions (N ₂ ⁺ ). Isotopic resolution for molecular ions of bovine ubiquitin (MW=8565 µ) also was achieved by using small ion populations.     

High throughput analysis of tryptophan metabolites in a complex matrix using capillary electrophoresis coupled to time-of-flight mass spectrometry

A capillary electrophoresis method for separation and detection with time-of-flight mass spectrometry is described for tryptophan metabolites in the kynurenic pathway. Tryptophan metabolites are usually difficult to detect with electrospray mass spectrometry since they have low surface activity and occur in low nanomolar to micromolar range in body fluids. Modification of the silica-wall with 1-(4-iodobutyl)4-aza-1-azoniabicyclo[2,2,2]octane iodide, also named M7C4I, has successfully been used to deactivate the fused silica wall and generate a stable reversed electroosmotic flow. Utilizing this advantage together with electrospray ionization time-of-flight mass spectrometry, which generates high resolution and fast acquisition monitoring of species, proved to be successful even for such a complex matrix like human cerebrospinal fluid.     

Macrophyllosaponin E: a novel compound from the roots of Astragalus. oleifolius

Macrophyllosaponin E, a novel cycloartane-type triterpene, has been isolated from the roots of Astragalus oleifolius. The structure elucidation of the compound was achieved by a combination of one- and two-dimensional NMR techniques [1H-1H-correlation spectroscopy (COSY), 1H-13C-heteronuclear multiple guantum correlation spectroscopy (HMQC), and 1H-13C-heteronuclear multiple-bond correlation spectroscopy (HMBC)l and high resolution electrospray ionization Fourier transformation mass spectrometry (HR-ESI-FT-MS).     

Characterization of complex assemblages of organic acids in geological samples by negative electrospray ionization mass spectrometry using a double‐focusing magnetic sector field mass spectrometer

Four different geological sample types (a crude oil, a crude oil asphaltene, a reservoir core extract and a reservoir core asphaltene) have been characterized by negative ionization electrospray mass spectrometry at low and high mass resolution using a double‐focusing magnetic sector field mass spectrometer. The mass range, shape of the spectra and the signal distribution of the acidic constituents as well as the average molecular weights, the total ion abundance and signal intensity in the spectra were compared for the different sample types. Nominal mass classes have been evaluated and Kendrick mass plots were generated in order to identify homologous series. For the crude oil sample, accurate mass assignments were made by high‐resolution double‐focusing magnetic sector field mass spectrometry (DFMSFMS) and were compared with those obtained by negative ion electrospray ionization (ESI) Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS). With both instrument types, compounds with the molecular composition C_nH_2n+zO₂, among which carboxylic acids predominated, were the main acidic compound class detectable in negative ESI mass spectra. Good agreement was achieved for the double bond class distribution and the carbon number distribution of the O₂ class. In addition, minor compound classes could be identified using FTICRMS. Copyright © 2010 John Wiley & Sons, Ltd.     

High resolution ICP-MS — a new concept for elemental mass spectrometry

Interfering molecular species are of major concern to the analyst currently using quadrupole based ICP-MS instrumentation. The recognized advantage and convenience offered by atmospheric plasma ionisation to multielement trace analysis can be significantly deteriorated by the limited resolving power of these analyzers. The result is poor sensitivity and lack of selectivity. With respect to sensitivity and resolution significant enhancement can be achieved by using magnetic sector based high resolution analyzers instead of quadrupoles. Unfortunately, up to now, commercially available HR-ICP-MS systems have been derived from complex instruments originally designed to meet the requirements of organic mass applications. Consequently, operation and performance of those systems expose the compromise which had to be made between an atmospheric plasma atomic ion source at high potential and an analyzer technology dedicated to molecular mass spectroscopy of organic compounds. At Finnigan MAT the first purpose designed high resolution ICP-MS has now been developed, which can be operated in high and low resolution mode at enhanced sensitivity. An innovative electric and magnetic field scanning strategy (SynchroScan) results in high on-peak duty cycles. Improved magnet technology offers high speed quadrupole style survey scans covering the full elemental mass range at nominal mass resolution. By extremely rapid peak switching, for example, all barium isotopes can be monitored in less than 100 ms with more than 90% on-peak detection efficiency. Examples are shown for computer controlled high and low resolution scan modes demonstrating the analytical performance of the new instrument concept. A comparison of detection limits achieved in low and high resolution mode is given.     

Multiresidue analysis of quinolones in water by ultra‐high perfomance liquid chromatography with tandem mass spectrometry using a simple and effective sample treatment

A rapid and simple analytical method has been developed for the determination of 19 quinolones in environmental water samples using ultra high performance liquid chromatography with tandem mass spectrometry. Chromatographic and detection conditions have been optimized and the separation was achieved in less than 4 min. The separation was carried out using a new‐generation column filled with superficially porous particles, resulting in lower backpressure and better resolution than totally porous particle columns. The quinolones were detected by electrospray ionization in positive mode using multiple‐reaction monitoring mode for acquisition. A sample treatment based on liquid–liquid extraction and phase separation via salting‐out was employed to achieve a fast and simple extraction that enables the multiresidue analysis. The method has been validated for an environmental well water sample from a mountain area. Very low limits of detection (between 10 and 90 ng/L) with relative standard deviations lower than 16.5% and recoveries higher than 73% were achieved. Moreover, well waters from different origins (mountain and coast areas and irrigated land) have been evaluated and similar results were obtained.     

Eurycomaoside: a new quassinoid-type glycoside from the roots of Eurycoma longifolia

A new C(19)-quassinoid-type glycoside has been isolated from the roots of Eurycoma longifolia. The structure elucidation of the compound was achieved by a combination of one- and two-dimensional NMR techniques, including (1)H-(1)H-correlation spectroscopy (COSY), (1)H-(13)C-heteronuclear correlation spectroscopy (HMQC), and (1)H-(13)C-heteronuclear multiple-bond correlation spectroscopy (HMBC), as well as high resolution electrospray ionization Fourier transformation mass spectrometry (HR-ESI-FT-MS) data. The C(1)-glycosidation site in the quassinoid framework is encountered for the first time.     

基于液相色谱-质谱技术的肾脏代谢组学分析方法研究

建立了一种基于高效液相色谱-超高分辨质谱技术的肾脏代谢组学分析方法。肾脏组织于液氮中研磨成粉后,采用甲基叔丁基醚/甲醇/水溶剂体系进行提取,分别得到强极性部分(下层)和弱极性部分(上层)提取物,依次采用HILIC亲水色谱柱和反相C18色谱柱进行梯度洗脱分离后,进行高分辨质谱分析。采用电喷雾离子源(ESI),正、负离子模式检测,扫描方式为全扫描,扫描范围为100~1000 Da,质量分辨率为120000。结果表明,该方法灵敏度高,专属性强,稳定性良好,可同时获取肾脏组织中的强极性和弱极性代谢物信息,可为肾脏疾病和药物肾毒性生物标志物的发现提供一种新的方法。     

A high-performance bio-tissue imaging method using air flow-assisted desorption electrospray ionization coupled with a high-resolution mass spectrometer

An air flow-assisted desorption electrospray ionization (AFADESI) MSI device was combined with a highresolution mass spectrometer to optimize the system parameters and achieve more accurate spatial distribution characteristics for compounds of interest while investigating bio-tissue sections. Finally, the parameter conditions that can provide optimal ionic intensity and enhanced resolution were confirmed.     

Analysis of process impurities in the basic drug SB-253149 using capillary electrophoresis and on-line mass spectrometric detection

Capillary electrophoresis with on-line electrospray ionisation mass spectrometry (CE/ESI-MS) has been used to identify process impurities in a batch of the anti-atherosclerotic drug, SB-253149. The impurities were separated from the main drug compound by capillary electrophoresis (CE) using an ammonium formate buffer at low pH in an untreated fused silica capillary. The CE method was initially developed using UV as the detection mode and then later structural elucidation work was achieved using an ion trap mass spectrometer. To maintain peak resolution and peak shape when the CE system was coupled to the mass spectrometer, a modified capillary cassette linked to a coaxial sheath flow electrospray ionisation (ESI) interface was used. By performing MS/MS experiments in conjunction with chemical knowledge of the reactivities of SB-253149, it was possible to propose molecular structures for impurities detected in the batch of SB-253149. The results from this study revealed that most of the process impurities in SB-253149 were dimeric derivatives of the parent molecule as well as trace levels of the starting material. This type of information was vital in process control and optimisation for the synthetic route for this drug.     

Reversed-phase liquid chromatographic method for the determination of selected room-temperature ionic liquid cations

The separation of selected 1-alkyl- and 1-aryl-3-methylimidazolium-based room temperature ionic liquid cations has been performed using reversed-phase high-performance liquid chromatography with electrospray ionization mass detection. The RP-HPLC method development started with the selection of a column taking into account especially the resolution of low molecular congeners of the selected group. Mobile phase composition was optimized for peak resolution, sensitivity and high reproducibility of retention values. The results of the method development were applied to the determination of exemplary ionic liquid species present in the medium used in cytotoxicity studies.     

Simultaneous separation of hydrophobic and hydrophilic peptides with a silica hydride stationary phase using aqueous normal phase conditions

The application of a silica hydride modified stationary phase with low organic loading has been investigated as a new type of chromatographic material suitable for the separation and analysis of peptides with electrospray ionization mass spectrometric detection. Retention maps were established to delineate the chromatographic characteristics of a series of peptides with physical properties ranging from strongly hydrophobic to very hydrophilic and encompassing a broad range of pI values (pI 5.5-9.4). The effects of low concentrations of two additives (formic acid and acetic acid) in the mobile phase were also investigated with respect to their contribution to separation selectivity and retention under comparable conditions. Significantly, strong retention of both the hydrophobic and the hydrophilic peptides was observed when high-organic low-aqueous mobile phases were employed, thus providing a new avenue to achieve high resolution peptide separations. For example, simultaneous separation of hydrophobic and hydrophilic peptides was achieved under aqueous normal phase (ANP) chromatographic conditions with linear gradient elution procedures in a single run, whilst further gradient optimization enabled improved peak efficiencies of the more strongly retained hydrophobic and hydrophilic peptides.     

Drug Pharmacokinetics Determined by Real‐Time Analysis of Mouse Breath

Noninvasive, real‐time pharmacokinetic (PK) monitoring of ketamine, propofol, and valproic acid, and their metabolites was achieved in mice, using secondary electrospray ionization and high‐resolution mass spectrometry. The PK profile of a drug influences its efficacy and toxicity because it determines exposure time and levels. The antidepressant and anaesthetic ketamine (Ket) and four Ket metabolites were studied in detail and their PK was simultaneously determined following application of different sub‐anaesthetic doses of Ket. Bioavailability after oral administration vs. intraperitoneal injection was also investigated. In contrast to conventional studies that require many animals to be sacrificed even for low‐resolution PK curves, this novel approach yields real‐time PK curves with a hitherto unmatched time resolution (10 s), and none of the animals has to be sacrificed. This thus represents a major step forward not only in animal welfare, but also major cost and time savings.     

High mass resolution breath analysis using secondary electrospray ionization mass spectrometry assisted by an ion funnel

In this study, we used secondary electrospray ionization mass spectrometry assisted by an ion funnel (IF) operating at ambient pressure to find compounds in the mass range of 100–500 m/z in online breath fingerprinting experiments. In low‐resolution experiments conducted on an ion trap instrument, we found that pyridine is present in breath of individuals long after drinking coffee. In high‐resolution experiments conducted on a Fourier transform ion cyclotron resonance, we found more than 30 compounds in the mass range of 100–500 m/z in analogous online breath experiments. More than a third of these compounds have molecular weights above 200 Daltons and have not been mentioned in previous studies. In low‐resolution experiments as well as experiments without the IF, these compounds could not be detected. Copyright © 2012 John Wiley & Sons, Ltd.     

Structural analysis of monoterpene glycosides extracted from Paeonia lactiflora Pall. using electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry and high-performance liquid chromatography/electrospray ionization tandem mass spectrometry

Paeoniflorin standard was first investigated by electrospray ionization Fourier transform ion cyclotron resonance tandem mass spectrometry (ESI-FTICR-MS/MS) using a sustained off-resonance irradiation (SORI) collision-induced dissociation (CID) method at high mass resolution. The experimental results demonstrated that the unambiguous elemental composition of product ions can be obtained at high mass resolution. Comparing MS/MS spectra and the experimental methods of hydrogen and deuterium exchange, the logical fragmentation pathways of paeoniflorin have been proposed. Then, the extracts of the traditional Chinese medicine Paeonia lactiflora Pall. were analyzed by high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI-MS/MS). By comparison with the ESI-FTICR-MS/MS data of paeoniflorin, the isomers paeoniflorin and albiflorin in Paeonia lactiflora Pall. have been identified using HPLC/MS with CID in an ion trap and in-source CID. Furthermore, using the characteristic fragmentation pathways, the retention times (t(R)) in HPLC and MS/MS spectra, the structures of three other kinds of monoterpene glycoside compounds have been identified on-line without time-consuming isolation. Thus an HPLC/ESI-MS method for the analysis of constituents in Paeonia lactiflora Pall. has been established.     

Unit resolution mass spectra of 112 kDa molecules with 3 Da accuracy

Previously, the highest molecular weight for a sample yielding a unit resolution mass spectrum was 67 kDa (marginal at 86 kDa), obtained with a 6. 2 T Fourier-transform mass spectrometer with electrospray ionization. Now with a 9. 4 T instrument, resolving power of 170,000 has been achieved for chondroitinase I (997 amino acids) and II (990 amino acids), making possible molecular weight assignments of 112,509 and 111,714, respectively, versus 112,508 and 111,713 calculated. Assisting these assignments was the noise reduction in the resolved isotopic peaks achieved by the time domain data sampling technique introduced by Senko, Marshall, and co-workers.     

Optimization of capillary electrophoresis conditions for coupling to a mass spectrometer via a sheathless interface

When optimizing a capillary electrophoresis/electrospray ionization mass spectrometry (CE/ESI-MS) system, consideration has to be given not only to the separation but also to the electrospray stability. Methods developed for CE/UV analysis of drugs and peptides were considered and modified to be suitable for a CE/MS system with a robust sheathless interface. Different concentrations of the organic modifiers acetonitrile, methanol and 2-propanol were used in the separation buffer. The type and concentrations of these modifiers were also compared with reference to electrospray stability, sensitivity and time of analysis. In addition, different ionic strengths in the buffers were evaluated with reference to electrospray stability. The repeatability was used for the estimation of electrospray stability. The degree to which these parameters influenced the separation and the ESI stability was studied using a nine-peptide standard mixture and the antibiotic drugs bacampicillin and ampicillin as test substances. The analysis time and resolution were used as measures of the efficiency of the separation. A time-of-flight MS analyzer was used since it has the potential advantages of becoming a better fit for integration of CE with MS owing to the speed and sensitivity of this mass analyzer. The detection limit, i.e. 1 microM, for bacampicillin was comparable to what could be achieved with CE/MS on a quadrupole instrument using selected ion monitoring and sheath flow ESI.     

Anisole, a new dopant for atmospheric pressure photoionization mass spectrometry of low proton affinity, low ionization energy compounds

Atmospheric pressure photoionization (APPI) is a novel method of ionization in liquid chromatography/mass spectrometry (LC/MS). It was originally developed in order to broaden the range of LC/MS ionizable compounds towards less polar compounds that cannot be analyzed by electrospray (ESI) and atmospheric pressure chemical ionization (APCI). Studies done thus far have shown that non-polar compounds that earlier were not ionizable in LC/MS can indeed be ionized by the use of APPI. However, the best ionization efficiency for low polarity samples has been achieved with low proton affinity (PA) solvents that are not suitable in reversed-phase LC (RP-LC). Here it is demonstrated that the signals for analytes with low proton affinities in acetonitrile can be increased 100-fold by using anisole as the dopant for APPI, which takes the sensitivity to the same level achieved in the analysis of high PA analytes.     

Binding constant determination of high-affinity protein-ligand complexes by electrospray ionization mass spectrometry and ligand competition

We describe an approach for the determination of binding constants for protein-ligand complexes with electrospray ionization mass spectrometry, based on the observation of unbound ligands competing for binding to a protein target. For the first time, dissociation constants lower than picomolar could be determined with good accuracy by electrospray ionization mass spectrometry. The presented methodology relies only on the determination of signal intensity ratios for free ligands in the low mass region. Therefore, all the advantages of measuring low masses with mass spectrometry, such as high resolution are preserved. By using a reference ligand with known binding affinity, the affinity of a second ligand can be determined. Since no noncovalently bound species are observed, assumptions about response factors are not necessary. The method is validated with ligands binding to avidin and applied to ligands binding to p38 mitogen-activated protein kinase.